title: The hydrophobic hinge region of rat DNA polymerase beta is critical for substrate binding pocket geometry.
abstract: The hydrophobic hinge of DNA polymerase beta facilitates closing and stabilization of the enzyme once the nucleotide substrate has bound. Alteration of the hydrophobic nature of the hinge by the introduction of a hydrophilic glutamine residue in place of isoleucine 260 results in an inaccurate polymerase. The kinetic basis of infidelity is lack of discrimination during the binding of substrate. The I260Q polymerase beta variant has lower affinity than wild type enzyme for the correct substrate and much higher affinity for the incorrect substrate. Our results demonstrate that the hinge is important for formation of the substrate binding pocket. Our results are also consistent with the interpretation that DNA polymerase beta discriminates the correct from incorrect substrate during the binding step.
authors.affiliation: Departments of Genetics, Therapeutic Radiology and Program in Microbiology, Yale University School of Medicine, New Haven, Connecticut 06520, USA.
authors.forename: Daniela
authors.initials: D
authors.lastname: Starcevic
authors.forename: Shibani
authors.initials: S
authors.lastname: Dalal
authors.forename: Joachim
authors.initials: J
authors.lastname: Jaeger
authors.forename: Joann B
authors.initials: JB
authors.lastname: Sweasy
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D004247:DNA / Q000737:chemistry / Q000378:metabolism; D019951:DNA Polymerase beta / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D005973:Glutamine; D007532:Isoleucine; D007700:Kinetics; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D051381:Rats; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D013379:Substrate Specificity
chemical_list: D011994:Recombinant Proteins; D007532:Isoleucine; D005973:Glutamine; D004247:DNA; D019951:DNA Polymerase beta