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Expression, purification and preliminary X-ray analysis of proliferating cell nuclear antigen from the archaeon Thermococcus thioreducens.

ABSTRACT: Proliferating cell nuclear antigen (PCNA) is a DNA sliding clamp which confers processivity on replicative DNA polymerases. PCNA also acts as a sliding platform that enables the association of many DNA-processing proteins with DNA in a non-sequence-specific manner. In this investigation, the PCNA from the hyperthermophilic archaeon Thermococcus thioreducens (TtPCNA) was cloned, overexpressed in Escherichia coli and purified to greater than 90% homogeneity. TtPCNA crystals were obtained by sitting-drop vapor-diffusion methods and the best ordered crystal diffracted to 1.86 A resolution using synchrotron radiation. The crystals belonged to the hexagonal space group P6(3), with unit-cell parameters a = b = 89.0, c = 62.8 A. Crystals of TtPCNA proved to be amenable to complete X-ray analysis and future structure determination.

SUBMITTER: Byrne-Steele ML

PROVIDER: S-EPMC2795597 | biostudies-literature | 2009 Sep

REPOSITORIES: biostudies-literature

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Expression, purification and preliminary X-ray analysis of proliferating cell nuclear antigen from the archaeon Thermococcus thioreducens.

Byrne-Steele Miranda L ML Ng Joseph D JD

Acta crystallographica. Section F, Structural biology and crystallization communications 20090822 Pt 9

Proliferating cell nuclear antigen (PCNA) is a DNA sliding clamp which confers processivity on replicative DNA polymerases. PCNA also acts as a sliding platform that enables the association of many DNA-processing proteins with DNA in a non-sequence-specific manner. In this investigation, the PCNA from the hyperthermophilic archaeon Thermococcus thioreducens (TtPCNA) was cloned, overexpressed in Escherichia coli and purified to greater than 90% homogeneity. TtPCNA crystals were obtained by sittin ...[more]

PMID: 19724129

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