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Crystal structure of SUMO-modified proliferating cell nuclear antigen.

ABSTRACT: Eukaryotic proliferating cell nuclear antigen (PCNA) is a replication accessory protein that functions in DNA replication, repair, and recombination. The various functions of PCNA are regulated by posttranslational modifications including mono-ubiquitylation, which promotes translesion synthesis, and sumoylation, which inhibits recombination. To understand how SUMO modification regulates PCNA, we generated a split SUMO-modified PCNA protein and showed that it supports cell viability and stimulates DNA polymerase ? activity. We then determined its X-ray crystal structure and found that SUMO occupies a position on the back face of the PCNA ring, which is distinct from the position occupied by ubiquitin in the structure of ubiquitin-modified PCNA. We propose that the back of PCNA has evolved to be a site of regulation that can be easily modified without disrupting ongoing reactions on the front of PCNA, such as normal DNA replication. Moreover, these modifications likely allow PCNA to function as a tool belt, whereby proteins can be recruited to the replication machinery via the back of PCNA and be held in reserve until needed.

SUBMITTER: Freudenthal BD

PROVIDER: S-EPMC3193175 | biostudies-literature | 2011 Feb

REPOSITORIES: biostudies-literature

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Crystal structure of SUMO-modified proliferating cell nuclear antigen.

Freudenthal Bret D BD Brogie John E JE Gakhar Lokesh L Kondratick Christine M CM Washington M Todd MT

Journal of molecular biology 20101215 1

Eukaryotic proliferating cell nuclear antigen (PCNA) is a replication accessory protein that functions in DNA replication, repair, and recombination. The various functions of PCNA are regulated by posttranslational modifications including mono-ubiquitylation, which promotes translesion synthesis, and sumoylation, which inhibits recombination. To understand how SUMO modification regulates PCNA, we generated a split SUMO-modified PCNA protein and showed that it supports cell viability and stimulat ...[more]

PMID: 21167178

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Project description:BackgroundThe high intracellular salt concentration required to maintain a halophilic lifestyle poses challenges to haloarchaeal proteins that must stay soluble, stable and functional in this extreme environment. Proliferating cell nuclear antigen (PCNA) is a fundamental protein involved in maintaining genome integrity, with roles in both DNA replication and repair. To investigate the halophilic adaptation of such a key protein we have crystallised and solved the structure of Haloferax volcanii PCNA (HvPCNA) to a resolution of 2.0 A.ResultsThe overall architecture of HvPCNA is very similar to other known PCNAs, which are highly structurally conserved. Three commonly observed adaptations in halophilic proteins are higher surface acidity, bound ions and increased numbers of intermolecular ion pairs (in oligomeric proteins). HvPCNA possesses the former two adaptations but not the latter, despite functioning as a homotrimer. Strikingly, the positive surface charge considered key to PCNA's role as a sliding clamp is dramatically reduced in the halophilic protein. Instead, bound cations within the solvation shell of HvPCNA may permit sliding along negatively charged DNA by reducing electrostatic repulsion effects.ConclusionThe extent to which individual proteins adapt to halophilic conditions varies, presumably due to their diverse characteristics and roles within the cell. The number of ion pairs observed in the HvPCNA monomer-monomer interface was unexpectedly low. This may reflect the fact that the trimer is intrinsically stable over a wide range of salt concentrations and therefore additional modifications for trimer maintenance in high salt conditions are not required. Halophilic proteins frequently bind anions and cations and in HvPCNA cation binding may compensate for the remarkable reduction in positive charge in the pore region, to facilitate functional interactions with DNA. In this way, HvPCNA may harness its environment as opposed to simply surviving in extreme halophilic conditions.

Dataset Information

Crystal structure of SUMO-modified proliferating cell nuclear antigen.

Publications

Crystal structure of SUMO-modified proliferating cell nuclear antigen.

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