title: Inactivation of NEIL2 DNA glycosylase by pyridoxal phosphate reveals a loop important for substrate binding.
abstract: Pyridoxal-5'-phosphate (PLP), in addition to its known metabolic functions, inactivates many DNA-dependent enzymes through conjugation to their critical amino groups. We have investigated the ability of PLP to inhibit bifunctional DNA repair glycosylases, which possess a catalytic amine. Of six enzymes tested, only endonuclease VIII-like protein 2 (NEIL2) was significantly inhibited by PLP. The inhibition was due to Schiff base formation between PLP and the enzyme. PLP-conjugated NEIL2 completely lost its ability to bind damaged DNA. Liquid chromatography/nanoelectrospray ionization tandem mass spectrometry of the products of proteolysis of pyridoxylated NEIL2 identified Lys50 as the site of modification. Thus, the beta2/beta3 loop where Lys50 is located in NEIL2 is important for DNA binding, presumably lies next to a phosphate-binding site, and may represent a target for regulation of the enzyme activity.
authors.affiliation: SB RAS Institute of Chemical Biology and Fundamental Medicine, 8 Lavrentieva Ave, Novosibirsk 630090, Russia.
authors.forename: Inga R
authors.initials: IR
authors.lastname: Grin
authors.forename: Robert A
authors.initials: RA
authors.lastname: Rieger
authors.forename: Dmitry O
authors.initials: DO
authors.lastname: Zharkov
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D045647:DNA Glycosylases / Q000037:antagonists & inhibitors / Q000737:chemistry*; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase / Q000037:antagonists & inhibitors / Q000737:chemistry*; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D017433:Protein Structure, Secondary; D011732:Pyridoxal Phosphate / Q000737:chemistry*; D012545:Schiff Bases / Q000737:chemistry; D021241:Spectrometry, Mass, Electrospray Ionization; D013379:Substrate Specificity; D053719:Tandem Mass Spectrometry
chemical_list: D012545:Schiff Bases; D011732:Pyridoxal Phosphate; D045647:DNA Glycosylases; C502613:Neil2 protein, mouse; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase; C473642:NEIL2 protein, human