Characterization of the human proliferating cell nuclear antigen physico-chemical properties: aspects of double trimer stability.
Its toroidal structure allows the proliferating cell nuclear antigen (PCNA) to wrap around and move along the DNA fiber, thereby dramatically increasing the processivity of DNA polymerization. PCNA is also involved in the regulation of a wide spectrum of other biological functions, including epigenetic inheritance. We have recently reported that mammalian PCNA forms a double trimer complex, which may be critically important in coordinating DNA replication and other cellular functions. To gain a better understanding of the stability of PCNA complexes, we characterized the physico-chemical properties of the PCNA structure by in vivo and in vitro approaches. The data obtained by gel filtration and nondenaturing gel electrophoresis of native PCNA molecules confirm our previous observations, obtained using formaldehyde crosslinking, in which PCNA exists in the cell as a double trimer. We have also found that optimal pH (pH 6.5-7.5) is critical for the stability of the PCNA structure. The presence or absence of ATP, dithiothreitol, and Mg2+ does not affect the stability of the PCNA trimer or double trimer. However, 0.02% SDS can effectively inhibit PCNA double trimer, but not single trimer, formation. Interestingly, glycerol and ammonium sulfate significantly destabilize both PCNA trimer and double trimer structures.
Naryzhny
Stanislav N
SN
Department of Research, Northeastern Ontario Regional Cancer Program at the Sudbury Regional Hospital, 41 Ramsey Lake Road, Sudbury, ON P3E 5J1, Canada.
Desouza
Leroi V
LV
Siu
K W Michael
KW
Lee
Hoyun
H
D002850:Chromatography, Gel; D019281:Dimerization; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D018809:Proliferating Cell Nuclear Antigen / Q000737:chemistry* / Q000302:isolation & purification; D020836:Protein Structure, Quaternary; D014407:Tumor Cells, Cultured
D018809:Proliferating Cell Nuclear Antigen