Structural study on the clamp PCNA-loading mechanism by the clamp loader RFC

2016 Fiscal Year Final Research Report

• Project/Area Number: 26440023
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Multi-year Fund
• Section: 一般
• Research Field: Structural biochemistry
• Research Institution: University of Yamanashi
• Principal Investigator: OYAMA Takuji 山梨大学, 総合研究部, 准教授 (60423133)
• Co-Investigator(Renkei-kenkyūsha): ISHINO Yoshizumi 九州大学, 農学研究院, 教授 (30346837) ; SHIRAI Tsuyoshi 長浜バイオ大学, バイオサイエンス学部, 教授 (00262890) ; MAYANAGI Kouta 九州大学, 生体防御医学研究所, 助教 (50418571)
• Project Period (FY): 2014-04-01 – 2017-03-31
• Keywords: DNA複製 / タンパク質DNA複合体 / 結晶構造解析

Outline of Final Research Achievements

To understand the detailed loading mechanism of the clamp PCNA onto DNA by the clamp-loader RFC in an ATP-dependent manner, we aimed (1) preparation of selenomethionine-labeled (SeMet) PCNA, (2) high resolution structure determination of RFC, and (3) crystallization and structure determination of the RFC-PCNA-DNA ternary complex. We determined the crystal structure of SeMet PCNA at 2.5 angstrom resolution. We established a novel preparation method of the highly purified recombinant RFC, but did not succeed in the high-resolution structure determination of the RFC. We performed crystallization screening of the RFC-PCNA-DNA ternary complex in a wide range of conditions, but obtained no crystals during this research project.

Free Research Field

構造生物化学