Article

ISSN 0006-2979, Biochemistry (Moscow), 2023, Vol. 88, No. 10, pp. 1528-1543 © The Author(s) 2023. This article is an open access publication.

Russian Text © The Author(s), 2023, published in Biokhimiya, 2023, Vol. 88, No. 10, pp. 1847-1866.

1528

REVIEW

Similarities and Differences in Photochemistry

of Type I and Type II Rhodopsins

Mikhail A. Ostrovsky

1,2

, Olga A. Smitienko

, Anastasia V. Bochenkova

and Tatiana B. Feldman

1,2,a

Faculty of Biology, Lomonosov Moscow State University, 119991 Moscow, Russia

Emanuel Institute of Biochemical Physics, 119334 Moscow, Russia

Faculty of Chemistry, Lomonosov Moscow State University, 119991 Moscow, Russia

e-mail: feldmantb@mail.ru

Received July 1, 2023

Revised July 20, 2023

Accepted August 12, 2023

Abstract— The diversity of the retinal-containing proteins (rhodopsins) in nature is extremely large. Fundamental similar-

ity of the structure and photochemical properties unites them into one family. However, there is still a debate about the

origin of retinal-containing proteins: divergent or convergent evolution? In this review, based on the results of our own

and literature data, a comparative analysis of the similarities and differences in the photoconversion of the rhodopsin

of typesI andII iscarried out. The results of experimental studies of the forward and reverse photoreactions of bacte-

riorhodopsin (type I) and visual rhodopsin (type II) in the femto- and picosecond time scale, photo-reversible reaction

of octopus rhodopsin (type II), photovoltaic reactions, as well as quantum chemical calculations of the forward photo-

reactions of bacteriorhodopsin and visual rhodopsin are presented. The issue of probable convergent evolution of type I

and type II rhodopsins is discussed.

DOI: 10.1134/S0006297923100097

Keywords: retinal-containing proteins, visual rhodopsin, bacteriorhodopsin, convergent evolution, photochemistry, femto-

second spectroscopy, quantum chemical calculations

Abbreviations: RCP,retinal-containing protein; RPSB,retinal protonated Schiff-base; BR,bacteriorhodopsin of Halobacterium

salinarum archaeon; Rh,bovine visual rhodopsin of Bostaurus; FC,Franck–Condon state; wave packet, a set of coherent excited

vibrational states; CI,conical intersection of potential energy surfaces; HOOP,hydrogen-out-of-plane vibrations; PES,potential

energy surface

* To whom correspondence should be addressed.

INTRODUCTION

Family of the retinal-containing proteins(RCPs) in-

clude three types of rhodopsin: microbial (typeI), ani-

mal (type II), and recently discovered heliorhodopsins

(type III) [1-7]. Despite the fact that their functions are

very diverse, fundamental similarity of the structure,

7-α-helical topography of the protein part and retinal as

a chromophore group, as well as of their photochemi-

cal and spectral properties is astounding. Obviously, this

raises the question of their evolutionary origin. Bearing

in mind conflicting opinions on their divergent [8-10] or

convergent [11-15] evolution, we consider the latter more

favorable. In other words, it has been suggested that there

is no common ancestor for all three types of rhodopsins.

And, if this is true, most likely, the pressure of external

factors (Darwin’s natural selection) and physiological

need resulted in such amazing similarity of such unrelat-

ed RCPs. In this regards comparison and understanding

of evolution of each type of rhodopsins seem to be an

independent topic that attract a lot of interest among the

researchers. The ever-increasing number of publications

on this topic testifies to this[3-5,11,16-18].

It became clear recently that the diversity of RCPs

is very large. At present they have been found in all do-

mains of life– bacteria, archaea, and eukaryotes, as well

PHOTOCHEMISTRY OF TYPE I AND II RHODOPSINS 1529

BIOCHEMISTRY (Moscow) Vol. 88 No. 10 2023

as in giant viruses. Type I rhodopsins are characteris-

tic for bacteria, archaea, viruses, and lower eukaryotes;

they have very diverse functions with main ones includ-

ing light-driven energy (ion pumps) and photo-infor-

mation (sensor rhodopsins, cation, and anion channels)

functions [6]. Type II rhodopsins are characteristic for

higher animals; in the majority of cases, they are repre-

sented by the specialized G-protein-coupled receptors,

which ensure mainly photo-information functions, pre-

dominantly vision function [1,2]. Type III rhodopsins

are widely available in live organisms from the same

domains of life as typeI rhodopsins, and they probably

perform photo-information functions [7].

Initially the term rhodopsin was referred to only pro-

teins mediating vision. The protein discovered in 1876

by Ferenz Boll was first named a vision compound

(Sehestoff), next, due to its color, “visual purple” (Seh-

purpur), and later “rhodopsin” from the Greek words

“rhodo”– pink and “opsis”– sight. The retinal-contain-

ing proton pump of halophilic archaeon (Halobacterium

salinarum) discovered by Walther Stoeckenius and Dieter

Oesterhelt almost 100 years later was named bacteri-

orhodopsin by analogy with the visual rhodopsin [19].

Atpresent the name rhodopsin is applied to all mem-

bers of typeI, II, and III RCPs. Their common features

are, first, the structure of apo-protein, opsin, with seven

transmembrane α-helices, and, second, cofactor (chro-

mophore)– retinal that absorbs light quanta. It becomes

more obvious with time that the 7-α-helical protein do-

main of RCPs is both very conserved and, at the same

time, it is characterized with very high plasticity. As to

retinal, it is, as a rule, covalently bound via the pro-

tonated Schiff-base (RPSB) with the lysine amino acid

residue of opsin in the seventh transmembrane α-he-

lix (TM7), and exist as all-trans (types I and III rho-

dopsins) or 11-cis (type II rhodopsins) isomeric forms.

It should be noted that the chromophore center is the

most conserved domain of opsin. Protein environment

of RPSB is crucial for both spectral tuning of the rho-

dopsin molecule, and for facilitating ultrafast and effi-

cient photochemical reaction of the chromophore isom-

erization, which is the basis for functioning of all RCPs.

Considering the role of protein environment of RPSB

in the chromophore center of opsin, it is acceptable

to consider the process of photoisomerization as cata-

lyzed by the protein. Furthermore, the issue of the nature

of protein–chromophore interactions (steric, electro-

static, hydrogen-bonding, and hydrophobic) is actively

investigated [20].

With regards to membrane topology of the protein

part of the molecule, it was established that in types I

and II rhodopsins N-end is facing outside the cell

and C-end– inside, while the opposite is observed in

typeIII rhodopsins with N-end facing inside the cell and

C-end – outside [4, 7, 21]. The reason and biological

significance of such arrangement of typeIII rhodopsin

is not yet clear. It is also surprizing that the “inverted”

topology of the protein part of the molecule has been

also observed in the olfactory G-protein-binding recep-

tors of insects[22].

Type I rhodopsins of bacteria and archaea, includ-

ing bacteriorhodopsin that performs the simplest pho-

tosynthesis, are among the most ancient proteins in the

biosphere, they emerged in prokaryotic cells around

3.8 billion years ago. Type I rhodopsins of single-cell

eukaryotes emerged 3.2billion years ago, while type II

rhodopsins of higher animals, including visual rhodop-

sin, appeared in multi-cellular eukaryotes less than 1bil-

lion years ago [5,11,18,23-25].

It has been suggested in the framework of the the-

ory of convergent evolution of rhodopsins that the

lysosomal cysteine transporters with 7-α-helical struc-

ture were predecessors of the microbial rhodopsins[15].

The cAMP-dependent G-protein-coupled receptors

with classic 7-α-helical structure, i.e., not containing

retinal receptors, are considered as predecessors of an-

imal rhodopsins [12-14], and retinal chromophore was

inserted in the chromophore centers of opsins later.

It has been suggested in one of the studies in the

framework of the theory of divergent evolution that, first,

type II rhodopsins originated from the cAMP receptors

(same as other G-protein-coupled receptors of classA),

from which later type I rhodopsins were generated via

horizontal gene transfer from eukaryotes to prokaryotes

[9]. In another work, based on optimization of electron

properties of the chromophore without considering clear

differences in amino acid sequences, the authors suggest

existence of the divergent pathway of evolution, but in

this case from type I to type II rhodopsins [8]. In one

other study the authors suggested their common origin

based on comparison of the structures of Na-binding

centers in the conserved amino acid region of the TM6

and on the functionally significant tilt of this helix in

the microbial rhodopsins and in the G-protein-coupled

receptors [10].

With regards to the recently discovered type III

rhodopsins, their origin remains obscure. According to

the suggestion of the authors of one study, heliorho-

dopsins originated from typeII eukaryotic rhodopsins,

which were next captured by the giant viruses and were

transferred to prokaryotic cells [16]. This implies, similar

to one of the hypotheses of the origin of typeI rhodo-

psins [9], unusual direction of evolution from eukaryotes

toprokaryotes.

In any case, the whole array of the data accumu-

lated so far allows to favor the notion on convergent in-

dependent evolution of all types of RCPs. In this review

similarities and differences in molecular mechanisms of

photochemical reaction in type I and type II rhodop-

sins are considered. Based on our own experimental

data and quantum-chemical calculations wepresent our

arguments in favor of the convergent evolution of RCPs.

OSTROVSKY et al.1530

BIOCHEMISTRY (Moscow) Vol. 88 No. 10 2023

COMPARISON OF THE PRIMARY

PHOTOREACTIONS INTYPEI

AND TYPEII RHODOPSINS

Rhodopsin functioning is based on the photochem-

ical reaction of isomerization of the RPSB chromo-

phore group (all-trans → 13-cis in typeI rhodopsins and

11-cis → all-trans in typeII rhodopsins) (Fig. 1a) [1, 2,

26-30]. Photoreaction occurs in the excited state and is

characterized with unique parameters, which are deter-

mined by both chemical properties of the chromophore

itself and effect of protein environment on the chromo-

phore [26, 31-33]. In the process, the energy of light

quantum is stored as the energy of the highly twisted

configuration of the isomerized RPSB in the chromo-

phore center[34]. Moreover, in the case of type I rho-

dopsins additional contribution to the energy storage is

provided by the change of the hydrogen bond network in

the RPSB region[27]. These processes occur on a fem-

to- and early-picosecond time scale [27]. Next, trans-

formation of the strained chromophore conformation

into a relaxed state occurs accompanied by the release

of stored energy, which, in turn, leads to re-arrangement

of the protein environment in the chromophore center.

Thisprocess eventually initiates global structural chang-

es of the entire protein part of the molecule required for

its functioning.

Mechanisms of photoinduced reactions of type I

and type II rhodopsins are presented in this section

of the review using the most studied representatives of

these two classes of RCPs, proton pump– bacteriorho-

dopsin of the H. salinarum archaeon (BR) and G-pro-

tein coupled receptor– bovine visual rhodopsin of Bos

taurus(Rh), as examples.

Stationary absorption spectra of BR and Rh are

shown in Fig.1b, which consist of α-, β-, and γ-bands

with the latter defined by the opsin absorption at 280 nm.

α- and β-bands are associated with the absorption of

RPSB as a part of the chromophore center. Depending

on the isomeric form, protonation state of the Schiff

base, and on peculiarities of the chromophore center

structure the spectral, photochemical, and a number

of other functionally important properties of the mol-

ecule change significantly[35]. Position of the α-band

absorption determines spectral range of rhodopsin func-

tioning (300-700nm)[2]. In the case of BR, maximum

of the α-band absorption is at 568 nm [Fig. 1b (1)], and

in the case of Rh – at 498 nm [Fig. 1b (2)]. Position

of the β-ionone ring significantly affects position of

the RPSB absorption maximum [36]. In BR the chro-

mophore group has planar trans-configuration of the

= C

and C

= C

bonds relative to the C

–C

single

bond [6-s-trans; Fig. 1a (1)], while in Rh it has cis-con-

figuration [6-s-cis; Fig. 1a (2)], which is not planar due

to steric hindrance causing decrease of the conjugation

length in the chromophore and shift of the Rh absorp-

tion maximum towards shorter wavelengths.

Forward photoreaction of type I and type II rhodop-

sins. Absorption of a quantum of light results in photo-

chemical reaction of RPSB isomerization and formation

of the primary photoproducts with spectral properties

different from the initial state. In this review our own

results on the dynamics of the primary processes of pho-

totransformation of BR and Rh obtained using method

of femtosecond absorption laser spectroscopy [37-46]

and supplemented with the data of quantum-chemical

calculations are presented [32,47-50].

Difference spectra and kinetic curves of the pho-

toinduced absorption of BR and Rh presented in Fig. 2

are produced with probing pulse delay time of up to 10ps.

The first photoinduced signals observed in the early fem-

tosecond time scale include absorption and stimulated

emission from the S

excited state [Fig. 2a (2and 3);

Fig. 2b (2)]. These signals are followed by absorption

of the first photoproduct in the ground electronic state

, and bleaching appears in the region of absorption

Fig. 1. Physicochemical characteristics of the chromophore group in type I and type II rhodopsins. a) Chemical structure of the rhodopsin

chromophore group– all-trans RPSB in typeI rhodopsins(1) and 6-s-cis–11-cis RPSB in typeII rhodopsins(2). Reactive bond is shown in bold.

b)Stationary absorption spectra of the suspension of purple membranes containing bacteriorhodopsin of the Halobacterium salinarum archaeon

(BR)(1) and of the detergent extract of the visual rhodopsin of Bos taurus(Rh)(2) normalized to α-band absorption.

PHOTOCHEMISTRY OF TYPE I AND II RHODOPSINS 1531

BIOCHEMISTRY (Moscow) Vol. 88 No. 10 2023

Fig. 2. Forward photoreactions of BR and Rh. a)Spectra of photoinduced absorption of BR recorded with delay times: –0.2(1), 0.05(2), 0.12(3),

0.5(4), 1(5), and 10(6) ps. b) Spectra of photoinduced absorption of Rh recorded with delay times: –0.2(1), 0.03(2), 0.1(3), 0.2(4), 0.8(5),

and 10(6)ps. Stationary absorption spectra of BR(a) and Rh(b) are presented with negative sign(7). c,d)Normalized kinetic curves of pho-

toinduced absorption of BR(1) and Rh(2) recorded at the absorption band of the excited state(c) with probing wavelengths of 470(BR) and

410(Rh)nm and in the absorption band of the photoreaction products(d) with probing wavelengths of 640(BR) and 580(Rh)nm. Kinetic curves

are shown in the linear scale of delay time up to 2ps and further in the logarithmic scale. The figure is adapted from[44] with permission.

of rhodopsin in the ground state [Fig. 2a (5), 600-

700 nm; Fig. 2b (4), 540-700 nm]. Within several pico-

seconds the absorption band of the first photoproduct

shifts slightly towards shorter wavelengths, which re-

flects formation of the next product as a result of the

processes of vibrational relaxation of the chromophore

group and its amino acid environment [Fig. 2a (6);

Fig. 2b (6)]. A fraction of the excited BR and Rh mol-

ecules returns back into the initial state with non-isom-

erized RPSB, which determined quantum yield of the

reaction.

Comparative analysis of the photoinduced absorp-

tion spectra of BR and Rh demonstrates differences in the

position of absorption bands and stimulated emission, as

well as in the time of formation of the primary products.

In the case of BR, the signals of the excited state(I

460

)

appear after delay time of 100 fs (Fig. 2a(2and3); 400-

540 nm and 700-880 nm) [42-44]. Within 1ps these sig-

nals practically completely disappear and are replaced

with the positive absorption signal of the first product

625

) [Fig. 2a (5)], which contains RPSB in 13-cis-con-

figuration. The next product (K

590

) is formed within a

picosecond time interval [Fig.2a(6)].

In the case of Rh, the signals from the excited

state(Rh

510

) appear within <30 fs (Fig. 2b (2); 410-480 nm

and 620-720 nm) [41, 42], which is significantly faster

than in the case of BR. At the delay time of 100 fs these

signals already disappear and are replaced with the pos-

itive signal of the first product (Photo

570

), which is com-

pletely formed by the time of 200 fs after absorption of

the light quantum [Fig. 2b (3and 4)] [37, 38, 41, 42].

The next product (Batho

535

) is formed within several

picoseconds [Fig.2b(5and6)].

Kinetic curves of the photoinduced absorption of

BR and Rh are presented in Fig. 2, which reflect for-

mation and decay of the excited state (I

460

and Rh

510

, re-

spectively; Fig. 2c) and formation of the first product of

photoreaction (J

625

and Photo

570

, respectively; Fig. 2d).

In the case of Rh, photoreaction proceeds much fast-

er (60 fs) in comparison with BR (480 fs) [40, 42-44],

which is in good agreement with the data reported by

Kochendoerfer and Mathies [51], Polli et al. [52], and

Johnson etal. [53, 54]. Time of formation of the sec-

ond product of the BR reaction (K

590

) is estimated as

1.8 ps, and of the second product of the Rh reaction

(Batho

535

)– as 2.2 ps [44]. Analysis of dynamics of the

OSTROVSKY et al.1532

BIOCHEMISTRY (Moscow) Vol. 88 No. 10 2023

Fig. 3. Kinetic schemes of elementary reaction of photoisomerization in type I rhodopsin– BR(a) and in typeII rhodopsin– Rh(b). Lifetimes

of the intermediate states are presented in the schemes as well as reactive(r) and non-reactive(nr) decay pathways of the excited state are marked.

Potential energy surfaces (S

andS

) of typeI rhodopsins exemplified by BR(c) and typeII rhodopsins exemplified by Rh(d) show the reaction

pathways of their forward (dark gray color) and reverse (gray color) photoreactions in femto- and pico-second time scale. FC,Franck–Condon

state; CI,conical intersection. Figure is adapted from[42] with permission.

excited state decay (both of Rh and BR) reveals that in

asmall fraction of the molecules (~4%) lifetime of the

excited state is much longer (2.4 ps), and its decay re-

sults in formation of only initial state of the RCP, hence,

this pathway has been termed non-reactive [44]. Exis-

tence of several pathways of the excited state decay, some

of which could be non-reactive, is, in general, typical

for type I rhodopsins [55-58], and in a lesser degree–

for type II rhodopsins [59]. This has been associated

with the split of the reaction pathway in the Frank–

Condon state (FC) or with the initial heterogeneity of

the protein part of the molecule [28, 60]. The latter sug-

gestion has been confirmed for the sodium pump KR2

of the Krokinobacter eikastus bacterium [49, 57] and

for the proton pumps proteorhodopsin and BR [58].

The ratio of the fraction of the excited molecules un-

dergoing isomerization and the fraction of the excited

molecules returning to the initial state via the reactive

and non-reactive pathways determines quantum yield

of isomerization, which is 0.64 for BR [61] and 0.65

for Rh [62]. Kinetic scheme and lifetimes of the ob-

served processes are presented in Fig. 3,a andb.

One of distinctive features of the photoreaction of

RPSB in rhodopsins and in solution is existence of the

vibrational component in its dynamics in the early stag-

es [29, 42, 53, 63, 64]. Phases and amplitudes of these

vibrations allowed to interpret their appearance as a re-

sult of non-stationary oscillatory motion in the excited

and ground states of the reagent and reaction products.

Contrary to the classic notions, formation of the prod-

uct in this type of photochemical reactions occurs via

coordinated motion of the molecule nuclei and ends

faster than the processes of vibrational relaxation and

vibrational dephasing in the excited state [65,66]. Such

reactions are termed coherent and could be described

as a motion of the wave packet (set of coherent excited

vibrational states) first down the S

potential energy sur-

face(PES) and next down the S

PES along the reaction

coordinate. Such fast S

→ S

transition is possible due

to existence of the multidimensional conical intersection

region(CI) of S

PESs, owing to which effective and

ultrafast transformation of light energy into chemical en-

ergy occurs [31,67].

The main differences in the dynamics of forward

photoreactions of BR and Rh are associated with the

structures of PESs of the S

, S

, and S

states of these

rhodopsins (Fig. 3, c and d), which is determined by

both isomeric form of RPSB, and by the effect of specif-

ic protein environment of the chromophore.

In the case of BR, similar to the other typeI rho-

dopsin, the forward photoreaction is described by the

3-state model including S

, S

, and S

states [26, 28,

55, 68], which postulates existence of a small barri-

er on the decay pathway of the S

state due to interac-

tion with the S

 PES, which significantly affects dy-

namics of the reaction (Fig. 3c). Excitation of the BR

results in formation of the FC state on the S

 PES,

which, as a result of the wave packet motion along

PHOTOCHEMISTRY OF TYPE I AND II RHODOPSINS 1533

BIOCHEMISTRY (Moscow) Vol. 88 No. 10 2023

Fig. 4. Optimized structures of reagents and primary products of BR and Rh– all-trans RPSB in BR

568

(a), 13-cis RPSB in K

590

(b), 11-cis RPSB in

498

(c) and all-trans RPSB in Batho

535

(d). Dihedral angles are shown by black lines with corresponding values.

all-symmetrical (C=C and C–C) vibrational modes of

RPSB, is transferred into the excited state I

460

within

40 fs. Next, the wave packet moves along the reactive hy-

drogen out-of-plane (HOOP) and torsional vibrational

modes and by overcoming a small barrier on the S

PES

reaches the CI region within the characteristic time of

480 fs. Inthe CI region the wave packed is divided into

two subpackets, with one of them is transferred to the

PES of the J

625

product, and the second one– to the

PES of the initial state BR

568

. As a result of overcoming

the barrier on the S

PES the wave packet loses its coher-

ent properties, which is obvious from the absence of clear-

ly pronounced oscillations in the time-resolved signals

of the product J

625

absorption [Fig. 2d(1)] [29, 42-44,

55,56]. This type of dynamics is termed diffusive[26].

In the case of Rh, same as for other typeII rhodop-

sins, forward photoreaction is described by the 2-state

model (S

andS

) (Fig. 3d) [2, 26, 28, 69]. As a result

of excitation of the Rh molecule a FC state is formed

on the S

 PES, which is transformed within less than

30 fs into the excited state Rh

510

as a result of motion

of the wave packet along the all-symmetric (C=C and

C–C) vibrational modes of RPSB, similarly to the case

of BR. Next, the wave packet moves along the out-of-

plane (HOOP and torsional) vibrational modes, reaches

the CI region within ≈60 fs, where it is divided into two

subpackets. The first subpacket moves along the S

PES

of Photo

570

, which is accompanied by the vibrational

relaxation of this product visualized by the clearly pro-

nounced oscillations of these kinetic curves (Fig. 2d),

while the second subpacket is transferred to the S

PES

of the initial state Rh

498

continuing its motion. Hence,

the S

→ S

transition of the wave packet in the process

of photoreaction of Rh occurs very fast, without devia-

tions and barriers thus preserving the significant portion

of coherence [37, 38, 42, 63]. Such type of dynamics is

termed ballistic or impulsive [26,70].

In order to explain differences in the dynamics of

photoinduced processes and specificity of the reaction

of photoisomerization in BR and Rh, quantum-chem-

ical calculations of the structures of these RCPs were

performed, and activity of the vibrational modes

during S

→ S

excitation and their link with the reac-

tive modes during non-radiative transition S

→ S

were

analyzed.

Optimized structures of the all-trans and 11-cis

RPSB in the ground electronic state in the protein envi-

ronment of BR and Rh, respectively, obtained with the

help of combined method of quantum and molecular

mechanics (QM/MM) are presented in Fig. 4 [48].

In the protein environment the chromophores

are found to have reactive bonds significantly twisted

(Fig. 4, a and c). On the other hand, these twists are

absent outside of the protein and the π-conjugated sys-

tem is planar with exception of the double bond in the

β-ionone ring. This bond, depending on the RPSB con-

figuration, is either in the conjugation plane of all other

five double bonds [6-s-trans; Fig. 1a (1)], or is out of

the conjugation plane [6-s-cis; Fig. 1a (2)]. It should be

noted that in the protein environment the out-of-conju-

gation-plane angle of the double bond in the β-ionone

ring in BR practically does not change, while in Rh it

increases significantly [48].

Hence, the protein environment indeed promotes

change of conformation and ‘twisting’ of the chromo-

phore group, which is due to its interaction with the

closest amino acid residues of the chromophore cen-

ter. This is mainly mediated by formation of hydrogen

bond between the protonated Schiff base and primary

counterion, which affects significantly torsion angle of

the reactive bond. Furthermore, attention should be also

paid to steric interaction in the area of β-ionone ring

of retinal. As expected, geometry of the primary pho-

toproducts was found to be even more twisted around

the reactive bonds due to their inability to relax in the

protein environment, which, on picosecond time scale,

remains more optimal for the initial isomers of RPSB

(Fig.4,bandd).

OSTROVSKY et al.1534

BIOCHEMISTRY (Moscow) Vol. 88 No. 10 2023

Fig. 5. Activity of the vibrational modes of RPSB during S

→S

transition in gas phase and in protein environment: all-trans RPSB in gas phase(a)

and inBR(b); 11-cis RPSB in gas phase(c) and inRh(d).

Interestingly enough, rotation barrier around the

single bond C

–C

does not exceed 3 kcal/mol in the gas

phase [50], which facilitates practically free transition

between the 6-s-trans and 6-s-cis conformations in the

RPSB. As was mentioned above, length of the π-con-

jugation chain is an important factor affecting position

of the absorption maximum wavelength. Intramolecular

rotation around the single C

–C

bond results in the ab-

normally wide absorption spectrum of the RPSB isomers

in the gas phase[71], while protein environment could

easily affect RPSB conformation in the region of β-ion-

one ring, and, depending on the value of dihedral angle

of the C

–C

bond, could control the length of π-con-

jugation in the chromophore thus adjusting its absorp-

tion to the certain range. Such mechanism of regulation

of photophysical properties of RPSB could be a basis for

spectral tuning in typeII rhodopsins. Noteworthy, posi-

tion of the β-ionone ring in BR does not change neither

in comparison with the gas phase, nor during transition

into the primary photoproducts, while its conformation

changes significantly in Rh. Hence, conformation of the

β-ionone ring is less significant in typeI rhodopsins in

comparison with type II rhodopsins.

Effects of twisting of the reactive bonds in different

RPSB isomers on the dynamics of isomerization in the

protein environment could be also illustrated by com-

paring it with the dynamics of photoresponse in the gas

phase. As was mentioned above, the RPSB isomers in

the isolated state have planar structure of the main part

of the π-conjugated system. It was shown with the help

of femtosecond spectroscopy and quantum-chemical

calculations that the relaxation dynamics of the electron-

ic excited state S

of the 11-cis RPSB occurs within the

sub-picosecond timeframe (400 fs) [32], which is com-

parable with the ultrafast times of photoisomerization

of the chromophore in Rh (50-100 fs) [40, 42, 51-54].

At the same time, specificity of the reaction and average

lifetimes of the excited state of the all-trans RPSB differ

significantly in the isolated state (3 ps)[32] and in the

protein environment in BR (≈500 fs) [29, 55, 56, 58].

It is important to mention that photoisomerization of

the planar all-trans-isomer of RPSB occurs slowly in

thegas phase, but selectively. The lowest barrier of isom-

erization in the S

state has been observed in the case

of the C

bond, but not in the case of the C

bond as in BR.

Hence, the effect of protein environment is a key

factor in the case of type I rhodopsins. While in the case

of type II rhodopsins, more advanced (from the point of

view of photochemical properties) 11-cis-isomer of RPSB

is used. Moreover, the reaction in Rh proceeds ultrafast,

which also implies significance of the effect of protein

environment of the chromophore center. Thisdifference

explains the clearly visible coherent mode of the primary

photochemical reaction in typeII rhodopsins.

The effect of conformation of the reactive bonds

in RPSB on the dynamics of photoisomerization has

been independently confirmed during investigation of

the chemically modified all-trans-chromophore twisted

around one of the double bonds. Change of the RPSB

conformation at the reactive bond results in the signif-

icant reduction of the photoisomerization barrier and

sub-picosecond times, which is typical for the reaction in

the protein environment[47].

Twisting of the reactive bond in the electronic

state S

not only changes the PES topography in the

state S

by decreasing the barrier of photoisomerization,

but also facilitates simultaneous excitation of certain

vibrational modes on transition S

→ S

[48, 49]. Inthe

process, excitation of both valent vibrations, localized

predominantly on the reactive double bond, and of

HOOP-vibrations at this bond occur. BR protein envi-

ronment in this case facilitates excitation of vibrations

of the C

bond, while in the case of Rh, protein

environment facilitates excitation of the C

bond.

Activity of the vibrational modes during photoexcitation

PHOTOCHEMISTRY OF TYPE I AND II RHODOPSINS 1535

BIOCHEMISTRY (Moscow) Vol. 88 No. 10 2023

could be estimated from the shift of minima of the PES

of the S

and S

states along these modes. The shifts

characterize changes of geometry of the chromophore in

the S

-state in comparison with the S

state. The larger is

the shift along certain vibration mode the more active is

vibration upon photoexcitation. The so-called Huang–

Rhys factor associated with the square of the shift along

the normal mode in a harmonic approximation is a use-

ful dimensionless parameter allowing evaluation of the

activity of the vibrational mode in electronic transition.

Values of the Huang–Rhys factor for the all-trans and

11-cis RPSB in the protein environment and in the gas

phase, calculated with the help of QM/MM method

using multiconfigurational quasidegenerate perturba-

tion theory of the second order, are presented in Fig. 5

[48, 49].

Two high-frequency valent vibrations of the bonds

and C

are active in the planar all-trans

and 11-cis RPSB upon excitation to the S

state in the

gas phase. In the case of BR, activity of the vibration-

al mode localized at the C

bond becomes signifi-

cantly higher in comparison with the gas phase, while

the valent vibration of the C

bond disappears en-

tirely. On the contrary, in the case of Rh, intensity of the

mode associated with vibrations of the C

bond in-

creases significantly, while intensity of the C

mode

decreases. Moreover, HOOP at the C

bond in Rh

also become active.

Hence, increase of twisting of the reactive bond

promotes increase of activity of the corresponding va-

lent vibration during photoexcitation. It should be men-

tioned that specificity of the photo-response of RPSB

and activity of the vibrations facilitating photoisomeri-

zation at the certain double bond (in this case valent and

HOOP vibrations) are more characteristic exactly for Rh.

This is in agreement with very high rate of the photo-

chemical reaction in this type II rhodopsin. The possi-

bility of excitation of various type of reactive modes in

the same phase, which promotes coherency of the reac-

tion in Rh, is also an important factor.

Hence, the results of quantum-chemical calcula-

tions provided explanation to the experimentally ob-

served difference in the dynamics of the photoinduced

processes in BR and Rh. The main factor affecting

these processes in both cases is protein environment of

the RPSB. Photoresponse of the chromophore group

in type I and type II rhodopsins becomes highly spe-

cific in comparison with the gas phase already at the

early times of the reaction resulting in excitation of cer-

tain vibrational modes, which facilitate isomerization

of the reactive bond. The 11-cis RPSB chromophore

in Rh is faster and more selective in comparison with

the all-trans RPSB in BR. Moreover, protein environ-

ment in Rh facilitates excitation of not only valent vibra-

tions of the reactive bond, but also of the out-of-plane

vibrations of hydrogen atoms at this bond, which directly

leads to the reaction of photoisomerization. It must be

also mentioned that conformation of the β-ionone ring

in the 6-s-cis-11-cis RPSB isomer, most likely, is an im-

portant factor in regulation of photophysical properties

of the chromophore group in typeII rhodopsins, which

is associated with the mechanism of spectral tuning.

It could be concluded that, most likely, the evo-

lutionary “younger” typeII rhodopsins have more ad-

vanced chromophore center in the opsin and isomeric

form of the chromophore, which allow more efficient

forward photochemical reaction.

Photochromism of type I and type II rhodopsins.

Rhodopsins exhibit photochromic properties [30, 34, 61,

72], i.e., have ability to realize phototransitions from the

intermediate products of the forward photoreaction back

to the initial state. This ability realized in nature in a

number of rhodopsins starting with the nanosecond time

scale serves for performing certain physiological func-

tions in such type I rhodopsins as sensory rhodopsin I

of H. salinarum [73], sensory rhodopsin of cyanobacte-

ria Anabaena sp. [29], and channel rhodopsin of the sin-

gle-cell alga Chlamydomonas reinhardtii[74], and in such

typeII rhodopsins as visual rhodopsins of invertebrates

[75-77] and closely related to them melanopsins of ver-

tebrates [78].

Despite the fact that at first stages of rhodopsin

phototransformation reverse phototransitions are practi-

cally not existing in nature, investigation of such ultrafast

photochromism under invitro conditions could provide

additional information on molecular mechanisms of the

photochemical reaction of these RCPs. In this review

the results of our studies investigating reverse photore-

actions of BR and Rh initiated in femto- and early pico-

second timeframes are presented [39,41,42].

Spectrum of photoinduced absorption of BR re-

corded with the delay time of 100 ps after the first pump

pulse with wavelength of 560 nm (pulse 1) that consists

of an absorption band of the product K

590

and bleach-

ing band of the initial state BR

568

is presented in Fig. 6

[Fig. 6a (1)]. The second pump pulse with wavelength

of 680 nm (pulse 2) and delay time of 5 ps excites K

590

molecules, part of which returns to BR

568

as a result of

reverse photoreaction (13-cis → all-trans RPSB). In the

process, dark formation of K

590

decreases and absorp-

tion of BR

568

increases [Fig. 6a (2)]. Quantum yield of

the reverse reaction of BR

568

formation from the prod-

uct K

590

is 0.81 [16], which is in agreement with the re-

sults reported by Kimetal.[61] and Balashovetal. [79].

Reverse phototransition could occur not only from the

product K

590

[Fig. 6a(inset:3and5 ps)], but also from

the product J

625

[Fig. 6a (inset:1 ps)] with approximately

equal efficiency [42].

In the case of Rh action of the pulse 2 (620 nm) that

follows with delay time after pulse 1 (500 nm) of 0.2-

3.8 ps also induces reverse photoreaction [Fig.6b][39,

41, 42, 46]. Depending on the pulse 2 delay the reverse

OSTROVSKY et al.1536

BIOCHEMISTRY (Moscow) Vol. 88 No. 10 2023

Fig. 6. Reverse photoreactions of BR and Rh. a,b)Spectra of photoinduced absorption of BR(a) and Rh(b) recorded with delay time of 100ps

after the action of one(1) and two(2) pump pulses with delay of the second pump pulse of 5 ps(a) and200 fs(b). In the spectral regions of

pump pulses experimental curves were completed using modeling (dashed lines). Inset: values of photoinduced absorption of BR(a) andRh(b)

recorded at probing wavelengths of 635nm(a) and560nm(b) before and after the second pump pulse depending on its delay time. c,d)Kinetic

curves of photoinduced absorption of BR(c) and Rh(d) recorded after the first pump pulse at probing wavelengths of 680 nm(c) and 620 nm(d).

Figure is adapted from[41,42] with permission.

phototransition occurs (all-trans → 11-cis RPSB) from

the Photo

570

product [Fig. 6b (inset:0.2-2 ps)] or from

the Batho

535

product [Fig. 6b (inset: 3-3.8 ps)]. Ef-

ficiency of the reverse photoreaction of Rh from the

Photo

570

product depends of the oscillation phase of

the time-resolved absorption signals of this product

[Fig. 6b(inset); Fig. 6d], and time of this photoreaction

is comparable or less than the time of forward photo-

reaction [41]. Quantum yield of the reverse photore-

action of Rh from the Batho

535

product is 0.15 [41, 42,

46], which is significantly less than the quantum yield

value (0.5) at 77 K reported previously by Suzuki and

Callender[80].

The reverse photochemical process in BR and Rh

initiated at early stages could be described as a motion of

the wave packet along the right branch of the S

PES of

the product of initial photochemical reaction towards the

CI region of the S

PES (Fig. 3,c andd). Theoretical

calculations for Rh demonstrated that this is the same

CI region, which participates in the forward photoreac-

tion [81]. Transition of the molecules from the excited

state S

to the S

PES via the CI region results in forma-

tion of the same states as in the forward reaction: BR

568

and J

625

– in the case of BR; Rh

498

and Photo

570

– in the

case of Rh. While efficiency of the reverse photoreaction

of BR practically does not depend on the delay of the

pulse 2 [Fig. 6a(inset)], for Rh it correlates directly with

the dynamics of the wave packet in thePhoto

570

product

[Fig. 6b (inset); Fig. 6d], which is associated with clearly

pronounced coherence of the forward photoreaction of

Rh. In this case, efficiency of the reverse photoreaction

correlates with the number of molecules of the Photo

570

product excited by the pulse2.

The rate of reverse photochemical reaction in rho-

dopsins is strongly affected by the particular isomeric

form of RPSB. Phototransition from trans- to cis-form

usually is slower than phototransition cis → trans, and

this is true for protein environment, gas phase, as well

as solution [26, 29, 32, 52, 57, 58, 82]. This difference is

due to the structures of S

, S

, and S

PES, as described

above using BR and Rh as examples [Fig. 3,c andd].

It was shown by Gaietal. [56] that, in the case of BR,

during excitation of the K

590

product the reverse photo-

chemical reaction from 13-cis- to all-trans-form of RPSB

occurs within the time period of ~100 fs, which is sig-

nificantly faster that the time of forward photoreaction

(480fs). It can be suggested that the reverse dependence

would be observed for Rh. However, as has been shown

experimentally for the Photo

570

product [41] and theoret-

ically for the Batho

535

product [81], time of the reverse

phototransition from all-trans- to 11-cis-form RPSB is

comparable with the time of forward phototransition

(60 fs) or less. It could be suggested that the rate (and

selectivity) of the reverse photochemical reaction of BR

and Rh is affected by the significant twist of the reactive

bond in the chromophore in their primary products by

the angle from –39° to –20° and from –36° to –34.8°,

respectively [31, 83]. This allows fast and barrier-less

PHOTOCHEMISTRY OF TYPE I AND II RHODOPSINS 1537

BIOCHEMISTRY (Moscow) Vol. 88 No. 10 2023

Fig. 7. Photochromic reaction of octopus rhodopsin. a)Difference spectra of two consecutive cycles of octopus rhodopsin transformations demon-

strating reversible conversion of rhodopsin into acidic metarhodopsin: metarhodopsin minus rhodopsin (1and3, after illumination with blue

light) and photoregeneration of rhodopsin: rhodopsin minus metarhodopsin (2and4, after illumination with red light). b)Photoreversibility

of octopus rhodopsin recorded at wavelength of maximum absorption of acidic metarhodopsin (528nm) under illumination with blue light(1),

red light(2), and in the dark(3). Figure is adapted from[76] with permission.

transition to the CI region even in the case of trans → cis

isomerization as in the reverse photoreaction of Rh.

It must be emphasized that while quantum yields of

the forward photoreaction in BR and Rh are practically

the same (≈0.65), quantum yields of their reverse photo-

reactions differ significantly (0.81 and 0.15, respectively).

According to the theoretical calculations [31, 81], quan-

tum yield of the product formation both in the forward

and reverse photoreactions of rhodopsins is determined

by the ratio of phases of HOOP and torsional vibration

modes of RPSB at the moment of the S

→ S

transition.

Initial population of the C=C and HOOP vibrational

modes at the reactive bond on excitation also plays an

important role in efficiency of the photoreaction, as was

mentioned above. In methanol solution the 13-cis → all-

trans and all-trans → 11-cis RPSB photoreactions proceed

with very close quantum yields (0.11 and 0.14, respec-

tively)[84]. Based on this, it could be suggested that the

significant difference in the quantum yields of the analo-

gous reverse reactions in BR and Rh is due to the effects

of protein environment at the chromophore center.

It could be mentioned that, from the function-

al point of view, the low quantum yield of the reverse

reaction in Rh in comparison with BR is an advantage

facilitating more reliable realization of the forward re-

action initiating the process of phototransduction. Light

absorption by type I rhodopsins serves primarily the en-

ergy conversion function, as in the case of BR, and re-

verse photoreactions likely cannot significantly affect

their efficiency. However, in the case of type II rho-

dopsins, which are G-protein-coupled receptors, 11-cis

RPSB acts as an effective ligand antagonist maintain-

ing low thermal “dark” noise of the photoreceptor cell.

On absorption of light, the isomerized all-trans RPSB

becomes a powerful ligand-agonist, and the activated Rh

initiates the process of phototransduction. Inthis case

the possibility of emergence of the reverse photoreaction

could significantly reduce efficiency of initiation of the

phototransduction process.

Hence, lower efficiency of the reverse photoreac-

tion of Rh, which increases reliability of the forward

photoreaction, could be considered as one of the ar-

guments in favor of selection in the course of evolution

of the 11-cis-isomer as a chromophore group in type II

rhodopsins. This indicates more advanced structure of

the chromophore center in Rh and evolutionary selec-

tion of such chromophore (11-cis RPSB), which allows

efficient phototransduction process. Obviously, under

real physiological conditions, the probability of the sec-

ond light quantum hitting the so short-lived intermedi-

ate of Rh photolysis as Batho

535

and more so Photo

570

is extremely low, however, potential possibility of reduc-

ing the probability of reverse photoreaction in the struc-

ture of Rh chromophore center exists.

At the same time, physiological significance of the

reverse photochemical reactions of type II rhodopsins

ininvertebrate animals from the late intermediate stag-

es of phototransformation is obvious. In the majority

of invertebrate animals this is one of the main ways for

regeneration (photoregeneration) of the visual pigment

[75, 77]. In particular, final product of the forward pho-

toreaction of the octopus rhodopsin is the so-called

acidic metarhodopsin, and of the reverse reaction –

again rhodopsin (Fig.7a) [76].

In the course of forward photoreaction, several in-

termediates with different absorption spectra are formed,

and in the course of reverse photoreaction– there are

only two states corresponding to conformational chang-

es first in the vicinity of chromophore, and, next, in

the entire protein, which return rhodopsin into initial

state. As we have demonstrated with the detergent ex-

tracts of the octopus rhodopsin, the photoreversible re-

action of typeII rhodopsin in invertebrates is extremely

stable, and under invitro conditions it can be repeated

multiple times for a significantly long period of time

(Fig.7b)[76].

Comparison of mechanisms of phototransformation

of type I and type II rhodopsins. Comparison of the

OSTROVSKY et al.1538

BIOCHEMISTRY (Moscow) Vol. 88 No. 10 2023

primary photoreactions of BR and Rh reveals both com-

mon and distinguishing features. Model kinetic schemes

(Fig.3,a andb) have much in common, and reflect the

suggestion that heterogeneity of the excited states of the

investigated rhodopsins is associated with heterogeneity

of their initial states. In the course of BR and Rh reac-

tions intermediates are formed that have similar proper-

ties: I

460

, J

625

, K

590

, and Rh

510

, Photo

570

, Batho

535

, respec-

tively. Elementary act of isomerization of RPSB proceeds

via transition through the CI of the S

PES, and time

of this transition is in femtosecond time scale. The first

products of photoinduced transformations of BR and Rh

with isomerized RPSB (J

625

and Photo

570

, respectively)

are formed in the ground(S

) state, and are vibrational-

ly excited precursors of the following products (K

590

and

Batho

535

, respectively) formed in the timeframe of sev-

eral picoseconds. At this stage isomerization of RPSB is

completed, and part of the light quantum energy is pre-

served in the strained structures of the products K

590

and

Batho

535

. Quantum yields of the forward photoreaction

of BR and Rh are practically the same (~0.65). Reaction

of photoisomerization of RPSB in BR and Rh, same as

in other rhodopsins, as well as in solution, is coherent

due to the inherent properties of the chromophore.

Dynamics and efficiency of the RPSB photoreac-

tion in rhodopsins depends on its isomeric form (through

the length of conjugated π-bonds) and on interactions

with the protein environment, which results not only in

the additional twisting of the chromophore, but also in

the strong electrostatic interactions with the counteri-

on. These factors affect the structure of the S

, S

, and

PES of the chromophore, by, for example, decreas-

ing barrier in the S

 PES of type I rhodopsins, which

significantly affects rate and coherence of the reaction.

Activity of certain vibrational modes also changes during

electronic transition S

→ S

. Selective and simultaneous

excitation of the vibrational modes associated with the

reactive modes of the non-radiative transition S

→ S

into the reaction products leads at the early stages of

photoinduced dynamics to high speed and high quantum

yield of the reaction of photoisomerization.

It can be concluded that interaction of RPSB with

opsin significantly increases the rate and quantum yield

of the photoreaction in comparison with the parameters

observed in gas phase and in solution, and selectivity of

the reaction reaches 100%. The finest regulation of these

parameter is achieved in type II rhodopsins, in which

the reaction has more pronounced coherence. The chro-

mophore–protein interactions also could be the cause

of heterogeneity of the initial state of RCPs, because

it is not observed in the gas phase [32].

Decrease of the length of π-conjugation in the

6-s-cis–11-cis RPSB in comparison with the all-trans-

isomer also leads to the shift of absorption maximum

of rhodopsin towards blue region of the spectrum and,

which is very important, to increase of the barrier of

thermal 11-cis → trans isomerization [26]. The first ef-

fect is actively used by typeII rhodopsins as one of the

factors of spectral tuning. The second factor is crucial

for reduction of the ‘dark’ noise of the photoreceptor

cell, which is also much facilitated by the limited volume

of the chromophore center [85]. At the same time, in

the case of BR and some other typeI rhodopsins, the

chromophore center is rather spacious and does not pre-

vent thermal trans → 13-cis isomerization occurring in

the dark.

All abovementioned features of the 11-cis RPSB are

extremely important for functioning of type II rhodop-

sins that perform photo-information function. This is,

likely, the reason why the 11-cis RPSB was select-

ed in the course of evolution as a chromophore group

of visual rhodopsins in both invertebrate and verte -

brate animals.

It should be emphasized that although the reaction

of photoisomerization is common for both type I and

type II rhodopsins, the final stages of the photoactivat-

ed processes differ significantly. In type I rhodopsins,

the 13-cis RPSB is subjected at one of the final stages

to thermal isomerization into the all-trans-form thus

closing the photoactivated processes into a photocy-

cle (Fig. 8a). In type II rhodopsins of the majority of

invertebrates, absorption of the second light quantum

Fig. 8. Schemes of type I rhodopsins photocycle exemplified by BR (a) and of photolysis of type II rhodopsins of vertebrates exemplified

by Rh(b).

PHOTOCHEMISTRY OF TYPE I AND II RHODOPSINS 1539

BIOCHEMISTRY (Moscow) Vol. 88 No. 10 2023

by one of the long-lived intermediate products of pho-

totransformation is required to close the cycle. The cy-

cle is not closed in type II rhodopsins of the vertebrates,

and the isomerized all-trans-retinal is released from the

chromophore center of the opsin, which is called pho-

tobleaching or rhodopsin photolysis (Fig.8b). In other

words, type II rhodopsin molecule of vertebrate ani-

mals– is a single-use molecule. For further function-

ing– rhodopsin regeneration– a new retinal molecule

in 11-cis conformation should be delivered to the chro-

mophore center (see details in the review by Ostrovsky

and Feldman[20]).

General similarities of the primary photochem-

ical reactions of type I and type II rhodopsins and

their significant differences in the following stages of

phototransformation have been manifested clearly in

their photoelectric reactions. In the end of 1970s and

beginning of 1980s using the method developed by

L. A. Drachev [86], we in collaborations with Drachev

and the Skulachev group compared in detail genera-

tion of photopotential for Rh on a photoreceptor disk

membrane from rods and for BR on a purple membrane

of haloarchaea [87-89]. As expected, both transition

all-trans → 13-cis in BR, and transition 11-cis → all-trans

in Rh were accompanied by the emergence of fast pho-

toelectric response. However, there was a significant

difference: while potential generation on the purple

membrane was accompanied also by the proton trans-

fer, which is essential for the photoenergetic function

of BR, in the case of Rh such transfer was absent. Itis

exactly the moment when physiological pathways of

type I and type II rhodopsins diverge. Absence of the

proton transfer across the photoreceptor disk mem-

brane was demonstrated experimentally in our later

study[90].

In the case of Rh, conformational changes in the

protein part of the molecule occur simultaneously with

the charge separation, which first results, at the me-

tarhodopsin II stage, in rapid deprotonation of RPSB

followed by slow release of the proton into external me-

dium[91]. Asa result of these conformational rearrange-

ments, Rh at the stage of metarhodopsinII acquires abil-

ity to interact with G-protein (transducin) and to initiate

the process of phototransduction [24, 92]. Based on its

characteristic times and polarity, the rapid component of

the Rh photoresponse is, in essence, the so-called early

receptor potential, which is observed during regular elec-

trophysiological separation from the retina [93].

While in the case of BR, with its photoactivated

processes connected into a cycle, stable electric po-

tential is generated on the purple membrane during

all period of illumination, in the case of Rh, which is

undergoing photolysis, rapid photoelectric response

irreversibly drops long before the light is switched

off, and there is no response to the repeated illu-

mination [88,89].

CONCLUSIONS

Hence, it can be concluded that the similarities of

type I and type II rhodopsins are associated with the

nature of their common chromophore group, retinal,

which, to a large extent, determines the mechanism

of photoreaction as a key stage of RCP functioning.

And the observed differences in the nature of photore-

actions of typeI and typeII rhodopsins most likely are

explained by the different isomeric form of the chromo-

phore selected in the course of evolution in accordance

with their functions. It can be suggested in the frame-

work of convergent evolution of type I and type II rho-

dopsins that the similar and at the same time plastic

7-α-helical protein structures of these two types evolved

independently driven by the need to optimize electronic

properties of their chromophore groups in the all-trans

and 11-cis isomeric forms.

Similarity of the structure and mechanism of ac-

tions of such complex molecular system as retinal-con-

taining transmembrane light-sensitive proteins is due

tothe pressure of natural selection with (must be em-

phasized) certain physicochemical limitations. The se-

lection itself was pursuing the most efficient function of

each type of rhodopsins.

It is absolutely clear that the mechanisms of evo-

lution of all three currently known types of RCPs [mi-

crobial (typeI), animal (typeII), and recently discov-

ered heliorhodopsins (typeIII)] require further detailed

investigation.

Contributions. M.A.O. concept and supervision of

the study; M.A.O., O.A.S., A.V.B., and T.B.F. writing

and editing of the paper.

Funding. This work was financially supported by the

Ministry of Science and Higher Education of the Rus-

sian Federation (agreement no.075-15-2020-795, inner

no.13.1902.21.0027).

Acknowledgments. The authors are grateful to

Dr. P. A. Kusochek and V. V. Logvinov for performing

part of quantum-chemical calculations. The calculations

were carried out using the HPC resources of the Lab-

oratory of Quantum Photodynamics (RSC Tornado)

provided through the Lomonosov Moscow State Uni-

versity Program of Development.

Ethics declarations. The authors declare no conflict

of interests in financial or any other sphere. This article

does not contain any studies with human participants

oranimals performed by any of the authors.

Open access. This article is licensed under a Cre-

ative Commons Attribution 4.0 International License,

which permits use, sharing, adaptation, distribution, and

reproduction in any medium or format, as long as you

give appropriate credit to the original author(s) and the

source, provide a link to the Creative Commons license,

and indicate if changes were made. The images or other

OSTROVSKY et al.1540

BIOCHEMISTRY (Moscow) Vol. 88 No. 10 2023

third-party material in this article are included in the ar-

ticle’s Creative Commons license, unless indicated oth-

erwise in a credit line to the material. If material is not

included in the article’s Creative Commons license and

your intended use is not permitted by statutory regula-

tion or exceeds the permitted use, you will need to obtain

permission directly from the copyright holder. To view

a copy of this license, visit http://creativecommons.org/

licenses/by/4.0/.

REFERENCES

1. Spudich, J. L., Yang, C.-S., Jung, K.-H., and Spudich,

E.N. (2000) Retinylidene proteins: structures and func-

tions from archaea to humans, Annu. Rev. Cell Dev. Biol.,

16, 365-392, doi:10.1146/annurev.cellbio.16.1.365.

2. Ernst, O. P., Lodowski, D. T., Elstner, M., Hegemann,P.,

Brown, L.S., and Kandori, H. (2014) Microbial and an-

imal rhodopsins: structures, functions, and molecular

mechanisms, Chem. Rev., 114, 126-163, doi: 10.1021/

cr4003769.

3. Nagata, T., and Inoue, K. (2021) Rhodopsins at a glance,

J.Cell Sci., 134, jcs258989, doi:10.1242/jcs.258989.

4. Rozenberg, A., Inoue, K., Kandori, H., and Béjà, O.

(2021) Microbial rhodopsins: The last two decades,

Annu. Rev. Microbiol., 75, 427-447, doi:10.1146/annurev-

micro-031721-020452.

5. Gordeliy, V., Kovalev, K., Bamberg, E., Rodriguez-

Valera, F., Zinovev, E., Zabelskii, D., Alekseev, A.,

Rosselli, R., Gushchin, I., and Okhrimenko, I. (2022)

Microbial rhodopsins, in Rhodopsin (Gordeliy, V., ed.)

Methods Mol. Biol., 2501, 1-52, Humana, New York, NY,

doi:10.1007/978-1-0716-2329-9_1.

6. Kandori, H. (2020) Biophysics of rhodopsins and opto-

genetics, Biophys. Rev., 12, 355-361, doi:10.1007/s12551-

020-00645-0.

7. Pushkarev, A., Inoue, K., Larom, S., Flores-Uribe, J.,

Singh, M., Konno, M., Tomida, S., Ito, S., Nakamu-

ra,R., Tsunoda, S.P., Philosof, A., Sharon, I., Yutin, N.,

Koonin, E.V., Kandori, H., and Béjà, O. (2018) A dis-

tinct abundant group of microbial rhodopsins discovered

using functional metagenomics, Nature, 558, 595-599,

doi:10.1038/s41586-018-0225-9.

8. Luk, H. L., Melaccio, F., Rinaldi, S., and Olivucci, M.

(2015) Molecular bases for the selection of the chromo-

phore of animal rhodopsins, Proc. Natl. Acad. Sci. USA,

112, 15297-15302, doi:10.1073/pnas.1510262112.

9. Mackin, K. A., Roy, R. A., and Theobald, D. L. (2014)

Anempirical test of convergent evolution in rhodopsins,

Mol. Biol. Evol., 31, 85-95, doi:10.1093/molbev/mst171.

10. Shalaeva, D. N., Galperin, M. Y., and Mulkidjanian,

A. Y. (2015) Eukaryotic G protein-coupled receptors as

descendants of prokaryotic sodium-translocating rho-

dopsins, Biol. Forward, 10, 63, doi: 10.1186/s13062-

015-0091-4.

11. Kojima, K., and Sudo, Y. (2023) Convergent evolution of

animal and microbial rhodopsins, RSC Adv., 13, 5367-

5381, doi:10.1039/d2ra07073a.

12. Krishnan, A., Almen, M. S., Fredriksson, R., and Schioth,

H. B. (2012) The origin of GPCRs: identification of mam-

malian like rhodopsin, adhesion, glutamate and frizzled GP-

CRs in fungi, PLoS One, 7, e29817, doi:10.1371/journal.

pone.0029817.

13. Feuda, R., Hamilton, S. C., McInerney, J. O., and

Pisani, D. (2012) Metazoan opsin evolution reveals a

simple route to animal vision, Proc. Natl. Acad. Sci. USA,

109, 18868-18872, doi:10.1073/pnas.1204609109.

14. Feuda, R., Menon, A. K., and Göpfert, M. C. (2022)

Rethinking opsins, Mol. Biol. Evol.,

39, 3, doi: 10.1093/

molbev/msac033.

15. Zhai, Y., Heijne, W. H., Smith, D. W., and Saier, M. H.,

Jr. (2001) Homologues of archaeal rhodopsins in plants,

animals and fungi: structural and functional predica-

tions for a putative fungal chaperone protein, Biochim.

Biophys. Acta, 1511, 206-223, doi: 10.1016/s0005-2736

(00)00389-8.

16. Bulzu, P.-A., Kavagutti, V. S., Andrei, A.-S., and

Ghai, R. (2022) The evolutionary kaleidoscope of rho-

dopsins, mSystems, 7, e00405-22, doi:10.1128/msystems.

00405-22.

17. Govorunova, E. G., Sineshchekov, O. A., and Spudich,

J. L. (2022) Emerging diversity of channelrhodopsins and

their structure-function relationships, Front. Cell. Neuros-

ci., 15, 800313, doi:10.3389/fncel.2021.800313.

18. Ostrovsky, M. A. (2017) Rhodopsin: evolution and

comparative physiology, Paleontol. J., 51, 562-572,

doi:10.1134/S0031030117050069.

19. Oesterhelt, D., and Stoeckenius, W. (1971) Rhodopsin-

like protein from the purple membrane of Halobacterium

halobium, Nat. New Biol., 233, 149-152, doi:10.1038/new-

bio233149a0.

20. Ostrovsky, M. A. and Feldman, T. B. (2012) Chemistry

and molecular physiology of vision: light-sensitive protein

rhodopsin, Russ. Chem. Rev., 81, 1071-1090, doi:10.1070/

RC2012v081n11ABEH004309.

21. Kovalev, K., Volkov, D., Astashkin, R., Alekseev, A.,

Gushchin, I., Haro-Moreno, J. M., Chizhov, I.,

Siletsky, S., Mamedov, M., Rogachev, A., Balandin, T.,

Borshchevskiy, V., Popov, A., Bourenkov, G., Bam-

berg,E., Rodriguez-Valera, F., Büldt, G., and Gordeliy,V.

(2020) High-resolution structural insights into the helior-

hodopsin family, Proc. Natl. Acad. Sci. USA, 117, 4131-

4141, doi:10.1073/pnas.1915888117.

22. Benton, R., Sachse, S., Michnick, S. W., and Vosshall,

L.B. (2006) Atypical membrane topology and heteromer-

ic function of Drosophila odorant receptors invivo, PLoS

Biol., 4, e20, doi:10.1371/journal.pbio.0040020.

23. Lamb, T. D., Collin, S. P., and Pugh, E. N., Jr. (2007)

Evolution of the vertebrate eye: Opsins, photorecep-

tors, retina and eye cup, Nat. Rev. Neurosci., 8, 960-976,

doi:10.1038/nrn2283.

PHOTOCHEMISTRY OF TYPE I AND II RHODOPSINS 1541

BIOCHEMISTRY (Moscow) Vol. 88 No. 10 2023

24. Hofmann, K. P., and Lamb, T. D. (2023) Rhodopsin,

light-sensor of vision, Prog. Retin. Eye Res., 93, 101116,

doi:10.1016/j.preteyeres.2022.101116.

25. Rozanov, A. Yu. (2009) Conditions of life on early earth

after 4.0 billion years ago, Problems of Origin of Life,

PINRAN, Moscow, pp.185-201.

26. Gozem, S., Luk, H. L., Schapiro, I., and Olivucci, M.

(2017) Theory and simulation of the ultrafast dou-

ble-bond isomerization of biological chromophores,

Chem. Rev., 117, 13502-13565, doi:10.1021/acs.chemrev.

7b00177.

27. Kandori, H. (2011) Protein-controlled ultrafast photo-

isomerization in rhodopsin and bacteriorhodopsin,

Supramolecular Photochemistry: Controlling Photo-

chemical Processes, Chapter 14, 571-595, doi: 10.1002/

9781118095300.ch14.

28. Diller, R. (2008) Primary reactions in retinal proteins,

in Ultrashort laser pulses in biology and medicine. Bio-

logical and Medical Physics, Biomedical Engineering,

(Braun M., Gilch P., Zinth W., eds), Springer, Berlin.

Heidelberg, Germany, Chapter 10, 243-277, doi:10.1007/

978-3-540-73566-3_10.

29. Wand, A., Gdor, I., Zhu, J., Sheves, M., and Ruhman, S.

(2013) Shedding new light on retinal protein photochem-

istry, Annu. Rev. Phys. Chem., 64, 437-458, doi:10.1146/

annurev-physchem-040412-110148.

30. Hampp, N. (2000) Bacteriorhodopsin as a photochromic

retinal protein for optical memories, Chem. Rev., 100,

1755-1776, doi:10.1021/cr980072x.

31. Schapiro, I., Melaccio, F., Laricheva, E. N., and Olivuc-

ci,M. (2011) Using the computer to understand the chem-

istry of conical intersections, Photochem. Photobiol. Sci.,

10,867-886, doi:10.1039/c0pp00290a.

32. Kiefer, H.V., Gruber, E., Langeland, J., Kusochek, P.A.,

Bochenkova, A.V., and Andersen, L. H. (2019) Intrinsic

photoisomerization dynamics of protonated Schiff-base

retinal, Nat. Commun., 10, 1210, doi:10.1038/s41467-019-

09225-7.

33. Agathangelou, D., Roy, P. P., del Carmen Marin, M.,

Ferre, N., Olivucci, M., Buckup, T., Léonard, J., and

Haacke, S. (2021) Sub-picosecond C=C bond photo-

isomerization: evidence for the role of excited state mix-

ing, Comptes Rendus Physique, 22, 111-138, doi: 10.5802/

crphys.41.

34. Birge, R. R., Cooper, T. M., Lawrence, A. F., Masthay,

M.B., Vasilakis, C., Fan Zhang, C., and Zidovetzki, R.

(1989) A spectroscopic, photocalorimetric, and theoreti-

cal investigation of the quantum efficiency of the primary

event in bacteriorhodopsin, J.Am. Chem. Soc., 111, 4063-

4074, doi:10.1021/ja00193a044.

35. Wang, W., Geiger, J. H., and Borhan, B. (2013) The pho-

tochemical determinants of color vision, BioEssays, 36,

65-74, doi:10.1002/bies.201300094.

36. Okada, T., Sugihara, M., Bondar, A.-N., Elstner, M.,

Entel, P., and Buss, V. (2004) The retinal conformation

and its environment in rhodopsin in light of a new 2.2 Å

crystal structure, J.Mol. Biol., 342, 571-583, doi:10.1016/

j.jmb.2004.07.044.

37. Smitienko, O. A., Shelaev, I. V., Gostev, F. E., Feldman,

T.B., Nadtochenko, V.A., Sarkisov, O.M., and Ostro-

vsky, M. A. (2008) Coherent processes in formation of

primary products of rhodopsin photolysis, Dokl. Biochem.

Biophys., 421, 194-198.

38. Smitienko, O. A., Mozgovaya, M. N., Shelaev, I. V.,

Gostev, F. E., Feldman, T. B., Nadtochenko, V.A., Sark-

isov, O. M., and Ostrovsky, M. A. (2010) Femtosecond

formation dynamics of primary photoproducts of visual

pigment rhodopsin, Biochemistry (Moscow), 75, 25-35,

doi:10.1134/s0006297910010049.

39. Mozgovaya, M. N., Smitienko, O. A., Shelaev, I. V.,

Gostev, F. E., Feldman, T. B., Nadtochenko, V. A.,

Sarkisov, O. M., and Ostrovsky, M.A. (2010) Photochr-

omism of visual pigment rhodopsin on the femtosecond

time scale: coherent control of retinal chromophore isom-

erization, Dokl. Biochem. Biophys., 435, 302-306.

40. Nadtochenko, V. A., Smitienko, O. A., Feldman, T. B.,

Mozgovaya, M. N., Shelaev, I. V., Gostev, F. E., Sarkisov,

O. M., and Ostrovsky, M. A. (2012) Conical intersection

participation in femtosecond dynamics of visual pigment

rhodopsin chromophore cis-trans photoisomerization,

Dokl. Biochem. Biophys., 446, 242-246.

41. Smitienko, O., Nadtochenko, V., Feldman, T., Balatska-

ya, M., Shelaev, I., Gostev, F., Sarkisov, O., and Ostro-

vsky, M. (2014) Femtosecond laser spectroscopy of the

rhodopsin photochromic reaction: A concept for ultra-

fast optical molecular switch creation (ultrafast reversible

photoreaction of rhodopsin), Molecules, 19, 18351-18366,

doi:10.3390/molecules191118351.

42. Feldman, T. B., Smitienko, O. A., Shelaev, I. V., Gostev,

F. E., Nekrasova, O. V., Dolgikh, D. A., Nadtochenko,

V. A., Kirpichnikov, M. P., and Ostrovsky, M. A. (2016)

Femtosecond spectroscopic study of photochromic re-

actions of bacteriorhodopsin and visual rhodopsin,

J. Photochem. Photobiol. B, 164, 296-305, doi: 10.1016/

j.jphotobiol.2016.09.041.

43. Smitienko, O. A., Nekrasova, O. V., Kudryavtsev, A.V.,

Yakovleva, M. A., Shelaev, I. V., Gostev, F. E., Dol-

gikh, D.A., Kolchugina, I. B., Nadtochenko, V. A., Kir-

pichnikov, M. P., Feldman, T. B., and Ostrovsky, M. A.

(2017) Femtosecond and picosecond dynamics of re-

combinant bacteriorhodopsin primary reactions com-

pared to the native protein in trimeric and monomeric

forms, Biochemistry (Moscow), 82, 490-500, doi:10.1134/

S0006297917040113.

44. Smitienko, O., Feldman, T., Petrovskaya, L., Nekraso-

va,O., Yakovleva, M., Shelaev, I., Gostev, F., Cherepan-

ov, D., Kolchugina, I., Dolgikh, D., Nadtochenko, V.,

Kirpichnikov, M., and Ostrovsky, M. (2021) Comparative

femtosecond spectroscopy of primary photoreactions of

Exiguobacterium sibiricum rhodopsin and Halobacteri-

um salinarum bacteriorhodopsin, J.Phys. Chem.B, 125,

995-1008, doi:10.1021/acs.jpcb.0c07763.

OSTROVSKY et al.1542

BIOCHEMISTRY (Moscow) Vol. 88 No. 10 2023

45. Medvedeva, A. S., Smitienko, O. A., Feldman, T. B.,

and Ostrovsky, M. A. (2020) Comparative study of pho-

tochemistry of microbial rhodopsins (type I) and ani-

mal’s rhodopsins (Type II), Zh. Evol. Biokhim. Fiziol., 56,

519-523, doi:10.31857/S0044452920070943.

46. Ostrovsky, M. A., and Nadtochenko, V. A. (2021) Fem-

tochemistry of rhodopsins, Russ. J. Phys. Chem. B,

15, 344-351, doi:10.1134/S1990793121020226.

47. Gruber, E., Kabylda, A. M., Nielsen, M. B., Rasmussen,

A. P., Teiwes, R., Kusochek, P. A., Bochenkova, A. V.,

and Andersen, L. H. (2022) Light driven ultrafast bioin-

spired molecular motors: Steering and accelerating pho-

toisomerization dynamics of retinal, J. Am. Chem. Soc.,

144, 69-73, doi:10.1021/jacs.1c10752.

48. Kusochek, P. A., Logvinov, V. V., and Bochenkova, A.V.

(2021) Role of the protein environment in photoisom-

erization of type I and type II rhodopsins: a theoreti-

cal perspective, Moscow Univ. Chem. Bull., 76, 407-416,

doi:10.3103/S0027131421060110.

49. Kusochek, P. A., Scherbinin, A. V., and Bochenkova,

A.V. (2021) Insights into the early-time excited-state dy-

namics of structurally inhomogeneous rhodopsin KR2,

J. Phys. Chem. Lett., 12, 8664-8671, doi: 10.1021/

acs.jpclett.1c02312.

50. Toker, Y., Svendsen, A., Bochenkova, A. V., and Ander-

sen, L. H. (2012) Probing the barrier for internal rotation

of the retinal chromophore, Angew. Chem. Int. Ed. Engl.,

51, 8757-8761, doi:10.1002/anie.201203746.

51. Kochendoerfer, G. G., and Mathies, R. A. (1996) Spon-

taneous emission study of the femtosecond isomerization

dynamics of rhodopsin, J.Phys. Chem., 100, 14526-14532,

doi:10.1021/jp960509+.

52. Polli, D., Altoè, P., Weingart, O., Spillane, K. M., Man-

zoni, C., Brida, D., Tomasello, G., Orlandi, G., Kuku-

ra, P., Mathies, R. A., Garavelli, M., and Cerullo, G.

(2010) Conical intersection dynamics of the primary pho-

toisomerization event in vision, Nature, 467, 440-443,

doi:10.1038/nature09346.

53. Johnson, P. J. M., Halpin, A., Morizumi, T., Prokhoren-

ko, V. I., Ernst, O.P., and Miller, R. J. D. (2015) Local

vibrational coherences drive the primary photochem-

istry of vision, Nat. Chem., 7, 980-986, doi: 10.1038/

nchem.2398.

54. Johnson, P. J. M., Farag, M. H., Halpin, A., Morizu-

mi,T., Prokhorenko, V. I., Knoester, J., Jansen, T. L.C.,

Ernst, O. P., and Miller, R. J. D. (2017) The primary

photochemistry of vision occurs at the molecular speed

limit, J. Phys. Chem. B, 121, 4040-4047, doi: 10.1021/

acs.jpcb.7b02329.

55. Hasson, K. C., Gai, F., and Anfinrud, P. A. (1996)

The photoisomerization of retinal in bacteriorhodop-

sin: experimental evidence for a three-state model, Proc.

Natl. Acad. Sci. USA, 93, 15124-15129, doi: 10.1073/

pnas.93.26.15124.

56. Gai, F., Hasson, K. C., McDonald, J. C., and Anfinrud,

P. A. (1998) Chemical dynamics in proteins: the pho-

toisomerization of retinal in bacteriorhodopsin, Science,

279, 1886-1891, doi:10.1126/science.279.5358.1886.

57. Tahara, S., Takeuchi, S., Abe-Yoshizumi, R., Inoue, K.,

Ohtani, H., Kandori, H., and Tahara, T. (2018) Origin of

the reactive and nonreactive excited states in the primary

reaction of rhodopsins: pH dependence of femtosecond

absorption of light-driven sodium ion pump rhodop-

sinKR2, J.Phys. Chem.B, 122, 4784-4792, doi:10.1021/

acs.jpcb.8b01934.

58. Chang, C.-F., Kuramochi, H., Singh, M., Abe-Yoshizu-

mi, R., Tsukuda, T., Kandori, H., and Tahara, T. (2022)

Aunified view on varied ultrafast dynamics of the prima-

ry process in microbial rhodopsins, Angew. Chem. Int. Ed.

Engl., 61, e202111930, doi:10.1002/anie.202111930.

59. Kandori, H., Futurani, Y., Nishimura, S., Shichida, Y.,

Chosrowjan, H., Shibata, Y., and Mataga, N. (2001) Ex-

cited-state dynamics of rhodopsin probed by femtosec-

ond fluorescence spectroscopy, Chem. Phys. Lett., 334,

271-276, doi:10.1016/S0009-2614(00)01457-3.

60. Kochendoerfer, G. G., and Mathies, R. A. (1995) Ultrafast

spectroscopy of rhodopsins– photochemistry at its best!

Isr.J. Chem., 35, 211-226, doi:10.1002/ijch.199500028.

61. Kim, J. E., Tauber, M. J., and Mathies, R. A. (2001)

Wavelength dependent cis-trans isomerization in vision,

Biochemistry, 40, 13774-13778, doi:10.1021/bi0116137.

62. Govindjee, R., Balashov, S. P., and Ebrey, T. G. (1990)

Quantum efficiency of the photochemical cycle of bac-

teriorhodopsin, Biophys. J., 58, 597-608, doi: 10.1016/

S0006-3495(90)82403-6.

63. Wang, Q., Shoenlein, R. W., Peteanu, L. A., Mathies,

R. A., and Shank, C. V. (1994) Vibrationally coherent

photochemistry in the femtosecond primary event of vi-

sion, Science, 266, 422-424, doi:10.1126/science.7939680.

64. Liebel, M., Schnedermann, C., Bassolino, G., Taylor,G.,

Watts, A., and Kukura, P. (2014) Direct observation of

the coherent nuclear response after the absorption of

a photon, Phys. Rev. Lett., 112, 238301, doi: 10.1103/

PhysRevLett.112.238301.

65. Zewail, A. H. (2000) Femtochemistry: atomic-scale dy-

namics of the chemical bond, J.Phys. Chem.A, 104, 5660-

5694, doi:10.1021/jp001460h.

66. Sarkisov, O. M., and Umanskii, S. Ya. (2001) Fem-

tochemistry, Russ. Chem. Rev., 70, 449-469, doi:10.1070/

RC2001v070n06ABEH000664.

67. Klessinger, M. (1995) Conical intersections and the mech-

anism of singlet photoreactions, Angew. Chem. Int. Ed.

Engl., 34, 549-551, doi:10.1002/anie.199505491.

68. Gozem, S., Johnson, P. J. M., Halpin, A., Luk, H. L.,

Morizumi, T., Prokhorenko, V. I., Ernst, O.P., Olivuc-

ci,M., and Miller, R. J. D. (2020) Excited-state vibron-

ic dynamics of bacteriorhodopsin from two-dimensional

electronic photon echo spectroscopy and multiconfigura-

tional quantum chemistry, J.Phys. Chem. Lett., 11, 3889-

3896, doi:10.1021/acs.jpclett.0c01063.

69. Rivalta, I., Nenov, A., Weingart, O., Cerullo, G., Garavel-

li, M., and Mukamel, S. (2014) Modelling time-resolved

PHOTOCHEMISTRY OF TYPE I AND II RHODOPSINS 1543

BIOCHEMISTRY (Moscow) Vol. 88 No. 10 2023

two-dimensional electronic spectroscopy of the primary

photoisomerization event in rhodopsin, J.Phys. Chem.B,

118, 8396-8405, doi:10.1021/jp502538m.

70. Hayashi, S., Tajkhorshid, E., and Schulten, K. (2009)

Photochemical reaction dynamics of the primary event of

vision studied by means of a hybrid molecular simulation,

Biophys.J., 96, 403-416, doi:10.1016/j.bpj.2008.09.049.

71. Rajput, J., Rahbek, D. B., Andersen, L. H., Hirshfeld, A.,

Sheves, M., Altoè, P., Orlandi, G., and Garavelli, M.

(2010) Probing and modeling the absorption of retinal pro-

tein chromophores invacuo, Angew. Chem. Intl. Ed. Engl.,

49, 1790-1793, doi:10.1002/anie.200905061.

72. Yoshizawa, T., and Wald, G. (1963) Pre-lumirhodop-

sin and the bleaching of visual pigments, Nature, 197,

1279-1286, doi:10.1038/1971279a0.

73. Takahashi, T., Mochizuki, Y., Kamo, N., and Kobatake,Y.

(1985) Evidence that the long-lifetime photointermedi-

ate of s-rhodopsin is a receptor for negative phototaxis in

Halobacterium halobium, Biochem. Biophys. Res. Commun.,

127, 99-105, doi:10.1016/s0006-291x(85)80131-5.

74. Bruun, S., Stoeppler, D., Keidel, A., Kuhlmann, U.,

Luck, M., Diehl, A., Geiger, M. A., Woodmansee, D.,

Trauner, D., Hegemann, P., Oschkinat, H., Hilde-

brandt,P., and Stehfest, K. (2015) Light-dark adaptation

of channelrhodopsin involves photoconversion between

the all-trans and 13-cis retinal isomers, Biochemistry,

54, 5389-5400, doi:10.1021/acs.biochem.5b00597.

75. Dixon, S. F., and Cooper, A. (1987) Quantum efficien-

cies of the reversible photoreaction of octopus rhodop-

sin, Photochem. Photobiol., 46, 115-119, doi: 10.1111 /

j.1751-1097.1987.tb04744.x.

76. Ostrovsky, M. A., and Weetall, H. H. (1998) Octopus rho-

dopsin photoreversibility of a crude extract from whole

retina over several weeks’ duration, Biosens. Bioelectron.,

13, 61-65, doi:10.1016/S0956-5663(97)00078-X.

77. Inoue, K., Tsuda, M., and Terazima, M. (2007) Photore-

verse reaction dynamics of octopus rhodopsin, Biophys.J.,

92, 3643-3651, doi:10.1529/biophysj.106.101741.

78. Koyanagi, M., and Terakita, A. (2008) Gq-coupled rho-

dopsin subfamily composed of invertebrate visual pigment

and melanopsin, Photochem. Photobiol., 84, 1024-1030,

doi:10.1111/j.1751-1097.2008.00369.x.

79. Balashov, S. P., Imasheva, E. S., Govindjee, R., and

Ebrey, T. G. (1991) Quantum yield ratio of the forward

and back light reactions of bacteriorhodopsin at low

temperature and photosteady-state concentration of the

bathoproduct K, Photochem. Photobiol., 54, 955-961,

doi:10.1111/j.1751-1097.1991.tb02116.x.

80. Suzuki, T., and Callender, R. H. (1981) Primary photo-

chemistry and photoisomerization of retinal at 77 degrees

K in cattle and squid rhodopsins, Biophys.J., 34, 261-270,

doi:10.1016/S0006-3495(81)84848-5.

81. Schapiro, I., Ryazantsev, M. N., Frutos, L. M., Ferré,N.,

Lindh, R., and Olivucci, M. (2011) The ultrafast pho-

toisomerizations of rhodopsin and bathorhodopsin are

modulated by bond length alternation and HOOP driven

electronic effects, J. Am. Chem. Soc., 133, 3354-3364,

doi:10.1021/ja1056196.

82. Schoenlein, R. W., Peteanu, L. A., Mathies, R. A., and

Shank, C. V. (1991) The first step in vision: Femtosec-

ond isomerization of rhodopsin, Science, 254, 412-415,

doi:10.1126/science.1925597.

83. Altoè, P., Cembran, A., Olivucci, M., and Garavelli, M.

(2010) Aborted double bicycle-pedal isomerization with

hydrogen bond breaking is the primary event of bacteri-

orhodopsin proton pumping, Proc. Natl. Acad. Sci. USA,

107, 20172-20177, doi:10.1073/pnas.1007000107.

84. Koyama, Y., Kubo, K., Komori, M., Yasuda, H., and

Mukai, Y. (1991) Effect of protonation on the isomeriza-

tion properties of n-butylamine Schiff base of isomeric reti-

nal as revealed by direct HPLC analyses: Selection of isom-

erization pathways by retinal proteins, Photochem. Photo-

biol., 54, 433-443, doi:10.1111/j.1751-1097.1991.tb02038.x.

85. Ikuta, T., Shihoya, W., Sugiura, M., Yoshida, K., Wa-

tari, M., Tokano, T., Yamashita, K., Katayama, K.,

Tsunoda, S. P., Uchihashi, T., Kandori, H., and Nure-

ki, O. (2020) Structural insights into the mechanism of

rhodopsin phosphodiesterase, Nat. Commun., 11, 5605,

doi:10.1038/s41467-020-19376-7.

86. Drachev, L. A., Kaulen, A. D., and Skulachev, V. P. (1977)

Temporal characteristics of bacteriorhodopsin as a molec-

ular biological generator of current, Mol. Biol. (Moscow),

11, 1377-1387.

87. Bol’shakov, V. I., Drachev, L. A., Kalamkarov, G. R.,

Kaulen, A. D., and Skulachev, V. P. (1979) Community

of properties of bacterial and visual rhodopsins: light ener-

gy conversion to electric potential difference, Dokl. Akad.

Nauk SSSR, 249, 1462-1466.

88. Drachev, L. A., Kalamkarov, G. R., Kaulen, A. D.,

Ostrovsky, M. A., and Skulachev, V.P. (1981) Fast stag-

es of photoelectric processes in biological membranes.

II. Visual rhodopsin, Eur. J. Biochem., 117, 471-481,

doi:10.1111/j.1432-1033.1981.tb06362.x.

89. Drachev, L. A., Kaulen, A. D., Khitrina, L. V., and

Skulachev, V. P. (1981) Fast stages of photoelectric pro-

cesses in biological membranes. I. Bacteriorhodopsin,

Eur.J. Biochem., 117, 461-470, doi:10.1111/j.1432-1033.

1981.tb06361.x.

90. Shevchenko, T. F., Kalamkarov, G. R., and Ostrovsky,

M.A. (1987) The lack of H

transfer across the photore-

ceptor membrane during rhodopsin photolysis [in Rus-

sian], Sensory Sistems, 1, 117-126.

91. Ostrovsky, M. A., Fedorovich, I. B., and Poliak, S. E.

(1968) Change in pH of the medium during illumination

of the retina and of a suspension of outer segments of pho-

toreceptors, Biofizika, 13, 338-339.

92. Kalamkarov, G. R., and Ostrovsky, M. A. (2002) Mo-

lecular mechanisms of visual reception, Moscow: Nauka,

pp.1-280.

93. Brindley, G. S., and Gardner-Medwin, A. R. (1966) The

origin of the early receptor potential of the retina, J.Physi-

ol., 182, 185-194, doi:10.1113/jphysiol.1966.sp007817.