Structural insights into the adaptation of proliferating cell nuclear antigen (PCNA) from Haloferax volcanii to a high-salt environment.
The sliding clamp proliferating cell nuclear antigen (PCNA) plays vital roles in many aspects of DNA replication and repair in eukaryotic cells and in archaea. Realising the full potential of archaea as a model for PCNA function requires a combination of biochemical and genetic approaches. In order to provide a platform for subsequent reverse genetic analysis, PCNA from the halophilic archaeon Haloferax volcanii was subjected to crystallographic analysis. The gene was cloned and expressed in Escherichia coli and the protein was purified by affinity chromatography and crystallized by the vapour-diffusion technique. The structure was determined by molecular replacement and refined at 3.5 A resolution to a final R factor of 23.7% (R(free) = 25%). PCNA from H. volcanii was found to be homotrimeric and to resemble other homotrimeric PCNA clamps but with several differences that appear to be associated with adaptation of the protein to the high intracellular salt concentrations found in H. volcanii cells.
Morgunova
Ekaterina
E
Karolinska Institutet, NOVUM, Centre of Structural Biochemistry, S-14157 Huddinge, Sweden. ekaterina.morgunova@ki.se
Gray
Fiona C
FC
Macneill
Stuart A
SA
Ladenstein
Rudolf
R
D000595:Amino Acid Sequence; D003001:Cloning, Molecular; D018360:Crystallography, X-Ray; D004926:Escherichia coli / Q000235:genetics; D019622:Haloferax volcanii / Q000737:chemistry* / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D018809:Proliferating Cell Nuclear Antigen / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D012492:Salts / Q000378:metabolism; D016415:Sequence Alignment
D018809:Proliferating Cell Nuclear Antigen; D012492:Salts