Characterization of the XRCC1-DNA ligase III complex in vitro and its absence from mutant hamster cells

摘要: The human DNA repair protein XRCC1 was overexpressed as a histidine-tagged polypeptide (denoted XRCC1-His) in Escherichia coli and purified milligram quantities by affinity chromatography. XRCC1-His complemented the mutant Chinese hamster ovary cell line EM9 when constitutively expressed from plasmid or introduced electroporation. directly interacted with ligase III vitro to form complex that resistant 2 M NaCl. equally well Bloom syndrome, HeLa MRC5 cells, indicating syndrome is normal this respect. Detection of on far Western blots radiolabelled indicated level reduced approximately 4-fold also EM-C11, second member complementation group. Decreased levels thus account for most 6-fold activity observed previously EM9. Immunodetection revealed decreased EM-C11 (> 10-fold), XRCC1-DNA much two CHO mutants.