Induced Fit in the Selection of Correct versus Incorrect Nucleotides by DNA Polymerase β

详细信息    查看全文

• 作者：Beth Moscato ; Monalisa Swain ; J. Patrick Loria
• 刊名：Biochemistry
• 出版年：2016
• 出版时间：January 19, 2016
• 年：2016
• 卷：55
• 期：2
• 页码：382-395
• 全文大小：798K
• ISSN：1520-4995

文摘

DNA polymerase β (Pol β) repairs single-nucleotide gapped DNA (sngDNA) by enzymatic incorporation of the Watson–Crick partner nucleotide at the gapped position opposite the templating nucleotide. The process by which the matching nucleotide is incorporated into a sngDNA sequence has been relatively well-characterized, but the process of discrimination from nucleotide misincorporation remains unclear. We report here NMR spectroscopic characterization of full-length, uniformly labeled Pol β in apo, sngDNA-bound binary, and ternary complexes containing matching and mismatching nucleotide. Our data indicate that, while binding of the correct nucleotide to the binary complex induces chemical shift changes consistent with the process of enzyme closure, the ternary Pol β complex containing a mismatching nucleotide exhibits no such changes and appears to remain in an open, unstable, binary-like conformation. Our findings support an induced-fit mechanism for polymerases in which a closed ternary complex can only be achieved in the presence of matching nucleotide.