Endonuclease Activity of MutL Protein of the <Emphasis Type="Italic">Rhodobacter sphaeroides</Emphasis> Mismatch Repair System |
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| Authors: | M V Monakhova A I Penkina A V Pavlova A M Lyaschuk V V Kucherenko A V Alexeevski V G Lunin P Friedhoff G Klug T S Oretskaya E A Kubareva |
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| Institution: | 1.Belozersky Institute of Physico–Chemical Biology,Lomonosov Moscow State University,Moscow,Russia;2.Chemistry Department,Lomonosov Moscow State University,Moscow,Russia;3.Gamaleya Research Institute of Epidemiology and Microbiology,Moscow,Russia;4.Bioengineering and Bioinformatics Department,Lomonosov Moscow State University,Moscow,Russia;5.Research Institute of System Development,Moscow,Russia;6.Institut für Biochemie,Justus–Liebig–Universit?t,Gieβen,Germany;7.Institut für Mikrobiologie und Molekularbiologie,Justus–Liebig–Universit?t,Gieβen,Germany |
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| Abstract: | 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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