Photostimulation of Intramolecular Electron Transfer in Nitrogenase

S. Yu. Druzhinin,¹ L. A. Syrtsova,^1,2 E. T. Rubtsova,¹ and N. I. Shkondina¹

¹Institute of Chemical Physics, Russian Academy of Sciences, Chernogolovka, Moscow Region, 142432 Russia; E-mail: syrtsova@icp.ac.ru

²To whom correspondence should be addressed.

Submitted June 27, 1996; revision submitted July 11, 1996.

Laser spectroscopy and the photochemical donor system eosin-NADH was used to study photoreduction of nitrogenase (Fe-protein_ox×Mo-Fe-protein)×MgATP [(Av2_ox×Av1)×MgATP] in the time range of 0-100 msec. Two initial stages of nitrogenase turnover were recorded: photoreduction of Av2 and electron transfer from Av2_red to Av1. The kinetics of photoreduction of Av2 was studied in solution and in the complex with Av1 at a concentration of eosin of up to 8×10^-5 M. The second-order rate constants for the two last reactions (1.06×10⁷ ± 1.06×10⁶ and 1.12×10⁷ ± 1.12×10⁶ M^-1×sec^-1, respectively) were determined, and processes accompanying these reactions were studied. In contrast to dithionite (a different electron donor), this photochemical donor system reduces Av2 in the complex with the same effectiveness as in the free state in solution. The rate constant for the reaction of electron transfer from Av2_red to Av1 was 1.0×10² sec^-1 at 20°C, which is equal to the rate of this reaction with Av2 reduced by dithionite. The data suggest that the rate-limiting step of nitrogenase turnover with the photochemical donor system is not related to photoreduction of Av2 in the complex with Av1 or the transfer of an electron from Av2_red to Av1.

KEY WORDS: Fe-protein, Mo-Fe-protein, eosin, time-resolved laser spectroscopy, flash photolysis.

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