The impact of single-nucleotide polymorphisms of human apurinic/apyrimidinic endonuclease 1 on specific DNA binding and catalysis

Human apurinic/apyrimidinic (AP) endonuclease APEI is a crucial enzyme of the base excision repair (BER) pathway, which is in charge of recognition and initiation of removal of AP-sites in DNA. It is known that some single-nucleotide polymorphism (SNP) variants of APEI have a reduced activity as compared to wild-type APEl. It has been hypothesized that genetic variation in APEI might be responsible for an increased risk of some types of cancer. In the present work, analysis of SNPs of the APE] gene was performed to select the set of variants having substitutions of amino acid residues on the surface of the enzyme globule and in the DNA-binding site, thereby affecting protein-protein interactions or the catalytic reaction, respectively. For seven APEI variants (R221C, N222H, R237A, G241R, M270T, R274Q and P311S), conformational dynamics and catalytic activities were examined. The conformational changes in the molecules of APEI variants and in a DNA substrate were recorded as fluorescence changes of Trp and 2-aminopurine residues, respectively, using the stopped-flow technique. The results made it possible to determine the kinetic mechanism underlying the interactions of the APEI variants with DNA substrates, to calculate the rate constants of the elementary stages, and to identify the stages of the process affected by mutation. (C) 2019 Elsevier B.V. and Societe Francaise de Biochimie et Biologie Moleculaire (SFBBM). All rights reserved.