ATP-dependent structural change of the eukaryotic clamp-loader protein, replication factor C.

Abstract:

:The eukaryotic DNA sliding clamp that keeps DNA polymerase engaged at a replication fork, called proliferating cell nuclear antigen (PCNA), is loaded onto the 3' ends of primer DNA through its interaction with a heteropentameric protein complex called replication factor C (RFC). The ATPase activity of RFC is necessary for formation of a functional PCNA clamp. In the present study, the sensitivity of RFC to partial proteolysis is used to show that addition of ATP, ATPgammaS, or ADP induces different structural changes in RFC. Direct observation by electron microscopy reveals that RFC has a closed two-finger structure called the U form in the absence of ATP. This is converted into a more open C form on addition of ATP. In contrast, the structural changes induced by ATPgammaS or ADP are limited. These results suggest that RFC adapts on opened configuration intermediately after ATP hydrolysis. We further observe that PCNA is held between the two fingers of RFC and propose that the RFC structure change we observe during ATP hydrolysis causes the attached PCNA to form its active ring-like clamp on DNA.

authors

Shiomi Y,Usukura J,Masamura Y,Takeyasu K,Nakayama Y,Obuse C,Yoshikawa H,Tsurimoto T

doi

10.1073/pnas.97.26.14127

keywords:

subject

Has Abstract

pub_date

2000-12-19 00:00:00

pages

14127-32

issue

26

eissn

0027-8424

issn

1091-6490

pii

97/26/14127

journal_volume

97

pub_type

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