[Front Cell Dev Biol. ]The Enigma of Substrate Recognition and Catalytic Efficiency of APE1-Like Enzymes

The Enigma of Substrate Recognition and Catalytic Efficiency of APE1-Like Enzymes Front Cell Dev Biol. 2021 Mar 26;9:617161. doi: 10.3389/fcell.2021.617161. eCollection 2021. Authors Anastasiia T Davletgildeeva 1 2 , Alexander A Ishchenko 3 , Murat Saparbaev 3 , Olga S Fedorova 1 , Nikita A Kuznetsov 1 Affiliations 1 Institute of Chemical Biology and Fundamental Medicine, Siberian Branch of the Russian Academy of Sciences, Novosibirsk, Russia. 2 Department of Natural Sciences, Novosibirsk State University, Novosibirsk, Russia. 3 Group "Mechanisms of DNA Repair and Carcinogenesis", Equipe Labellisée LIGUE 2016, CNRS UMR 9019, Université Paris-Saclay, Villejuif, France. PMID: 33842455 PMCID: PMC8033172 DOI: 10.3389/fcell.2021.617161 Abstract Despite significant achievements in the elucidation of the nature of protein-DNA contacts that control the specificity of nucleotide incision repair (NIR) by apurinic/apyrimidinic (AP) endonucleases, the question on how a given nucleotide is accommodated by the active site of the enzyme remains unanswered. Therefore, the main purpose of our study was to compare kinetics of conformational changes of three homologous APE1-like endonucleases (insect Drosophila melanogaster Rrp1, amphibian Xenopus laevis xAPE1, and fish Danio rerio zAPE1) during their interaction with various damaged DNA substrates, i.e., DNA containing an F-site (an uncleavable by DNA-glycosylases analog of an AP-site), 1,N6-ethenoadenosine (εA), 5,6-dihydrouridine (DHU), uridine (U), or the α-anomer of adenosine (αA). Pre-steady-state analysis of fluorescence time courses obtained for the interaction of the APE1-like enzymes with DNA substrates containing various lesions allowed us to outline a model of substrate recognition by this class of enzymes. It was found that the differences in rates of DNA substrates' binding do not lead to significant differences in the cleavage efficiency of DNA containing a damaged base. The results suggest that the formation of enzyme-substrate complexes is not the key factor that limits enzyme turnover; the mechanisms of damage recognition and cleavage efficacy are related to fine conformational tuning inside the active site. Keywords: DNA repair; abasic site; apurinic/apyrimidinic endonuclease; pre-steady state kinetics; target nucleotide recognition. Copyright © 2021 Davletgildeeva, Ishchenko, Saparbaev, Fedorova and Kuznetsov. Conflict of interest statement The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.