Endonuclease Activity of MutL Protein of the Rhodobacter sphaeroides Mismatch Repair System

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M. V. Monakhova¹, A. I. Penkina², A. V. Pavlova², A. M. Lyaschuk³, V. V. Kucherenko⁴, A. V. Alexeevski^1,5, V. G. Lunin³, P. Friedhoff⁶, G. Klug⁷, T. S. Oretskaya², and E. A. Kubareva^1*

¹Belozersky Institute of Physico-Chemical Biology, Lomonosov Moscow State University, 119991 Moscow, Russia; E-mail: kubareva@belozersky.msu.ru

²Lomonosov Moscow State University, Chemistry Department, 119991 Moscow, Russia

³Gamaleya Research Institute of Epidemiology and Microbiology, 123098 Moscow, Russia

⁴Lomonosov Moscow State University, Bioengineering and Bioinformatics Department, 119991 Moscow, Russia

⁵Research Institute of System Development, 117218 Moscow, Russia

⁶Institut für Biochemie, Justus-Liebig-Universität, 35392, Gießen, Germany; E-mail: Sekretariat.Biochemie@chemie.bio.uni-giessen.de

⁷Institut für Mikrobiologie und Molekularbiologie, Justus-Liebig-Universität, 35392, Gießen, Germany; E-mail: gabriele.klug@mikro.bio.uni-giessen.de

^* To whom correspondence should be addressed.

Received September 12, 2017; Revision received November 11, 2017

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We have purified the MutL protein from Rhodobacter sphaeroides mismatch repair system (rsMutL) for the first time. rsMutL demonstrated endonuclease activity in vitro, as predicted by bioinformatics analysis. Based on the alignment of 1483 sequences of bacterial MutL homologs with presumed endonuclease activity, conserved functional motifs and amino acid residues in the rsMutL sequence were identified: five motifs comprising the catalytic site responsible for DNA cleavage were found in the C-terminal domain; seven conserved motifs involved in ATP binding and hydrolysis and specific to the GHKL family of ATPases were found in the N-terminal domain. rsMutL demonstrated the highest activity in the presence of Mn²⁺. The extent of plasmid DNA hydrolysis declined in the row Mn²⁺ > Co²⁺ > Mg²⁺ > Cd²⁺; Ni²⁺ and Ca²⁺ did not activate rsMutL. Divalent zinc ions inhibited rsMutL endonuclease activity in the presence of Mn²⁺ excess. ATP also suppressed plasmid DNA hydrolysis by rsMutL. Analysis of amino acid sequences and biochemical properties of five studied bacterial MutL homologs with endonuclease activity revealed that rsMutL resembles the MutL proteins from Neisseria gonorrhoeae and Pseudomonas aeruginosa.

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KEY WORDS: DNA repair, mismatch, MutL protein

DOI: 10.1134/S0006297918030082

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