Involvement of subcomplexes of 32 and 14 kDa subunits in RPA's DNA binding activity through redox change.
The eukaryotic replication protein A (RPA) is a heterotrimeric protein complex. It consists of 70, 32, and 14 kDa subunits that are involved in DNA replication, repair, and genetic recombination. RPA is a 4-cysteine type zinc-finger protein. RPA's zinc-finger domain is not essential for DNA binding activity, but it is involved in the regulation of RPA's DNA binding activity through reduction-oxidation (redox). In this study, we show that yeast RPA's ssDNA binding activity is regulated by redox potential through its subcomplexes of 32 and 14 kDa subunits. In contrast, the subunits' complex, RPA70, formed a stable complex with ssDNA, even under non-reducing conditions. The addition of DTT and H202 had no effect on its DNA binding activity. In RPA70, since the addition of the subcomplexes of the 32 and 14 kDa subunits, it restored the modulating ssDNA binding activity to native RPA's DNA binding activity. These results suggest that the subcomplexes of the 32 and 14 kDa subunits may be involved in the modulating RPA's DNA binding activity through redox change. These studies, therefore, show the novel structure and function relationship of a multiprotein complex in that the role of a specific domain (or one subunit) is regulated by the other subunits.
Department of Chemistry and Chemistry Institute for Functional Materials, Pusan National University, Busan, Korea.
Andre
A
Kim
Jang-Su
JS
Park
D004247:DNA / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D024202:Electrophoretic Mobility Shift Assay; D005656:Fungal Proteins / Q000378:metabolism; D008970:Molecular Weight; D010084:Oxidation-Reduction; D011485:Protein Binding; D021122:Protein Subunits / Q000378:metabolism; D051716:Replication Protein A
D004268:DNA-Binding Proteins; D005656:Fungal Proteins; D021122:Protein Subunits; D051716:Replication Protein A; D004247:DNA