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Modulation of the Pyrococcus abyssi NucS endonuclease activity by replication clamp at functional and structural levels.


ABSTRACT: Pyrococcus abyssi NucS is the founding member of a new family of structure-specific DNA endonucleases that interact with the replication clamp proliferating cell nuclear antigen (PCNA). Using a combination of small angle x-ray scattering and surface plasmon resonance analyses, we demonstrate the formation of a stable complex in solution, in which one molecule of the PabNucS homodimer binds to the outside surface of the PabPCNA homotrimer. Using fluorescent labels, PCNA is shown to increase the binding affinity of NucS toward single-strand/double-strand junctions on 5' and 3' flaps, as well as to modulate the cleavage specificity on the branched DNA structures. Our results indicate that the presence of a single major contact between the PabNucS and PabPCNA proteins, together with the complex-induced DNA bending, facilitate conformational flexibility required for specific cleavage at the single-strand/double-strand DNA junction.

SUBMITTER: Creze C 

PROVIDER: S-EPMC3346078 | biostudies-literature | 2012 May

REPOSITORIES: biostudies-literature

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Modulation of the Pyrococcus abyssi NucS endonuclease activity by replication clamp at functional and structural levels.

Creze Christophe C   Ligabue Alessio A   Laurent Sébastien S   Lestini Roxane R   Laptenok Sergey P SP   Khun Joelle J   Vos Marten H MH   Czjzek Mirjam M   Myllykallio Hannu H   Flament Didier D  

The Journal of biological chemistry 20120319 19


Pyrococcus abyssi NucS is the founding member of a new family of structure-specific DNA endonucleases that interact with the replication clamp proliferating cell nuclear antigen (PCNA). Using a combination of small angle x-ray scattering and surface plasmon resonance analyses, we demonstrate the formation of a stable complex in solution, in which one molecule of the PabNucS homodimer binds to the outside surface of the PabPCNA homotrimer. Using fluorescent labels, PCNA is shown to increase the b  ...[more]

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