ns and Thermus aquaticus consist of a homodimer of the peptide containing two OB-folds. eukaryotic RPA is a stable heterotrimer, composed of 70, 32, and 14 kDa proteins. RPA70 contains two tandem repeats of an OB-fold, which are responsible for the major interaction with a single-stranded DNA in its central region. The N-terminal and C-terminal regions of RPA70 mediate interactions with RPA32 and also with many cellular or viral proteins. RPA32 contains an OB-fold in the central region, and the C-terminal region interacts with other RPA subunits and various cellular proteins. RPA14 also contains an OB-fold. eukaryotic RPA interacts with the SV40 T-antigen and the DNA polymerase?-Primase complex, and thus forms part of the initiation complex at the replication origin. The RPA also stimulates Pol?-Primase activity and PCNA-dependent Pol? activity. The RPAs from M. jannaschii and M. thermautotrophicus were reported in 1998, as the first archaeal single-stranded DNA binding proteins. These proteins share amino acid sequence similarity with the eukaryotic RPA70, and contain four or five repeated OB-fold and one zincfinger motif. M. jannaschii RPA exists as a monomer in solution, and has single-strand DNA binding activity. On the other hand, P. furiosus RPA forms a complex consisting of three distinct subunits, RPA41, RPA32, and RPA14, similar to the eukaryotic RPA. The P. furiosus RPA strikingly stimulates the RadA-promoted strand-exchange reaction in vitro . the euryarchaeal organisms have a eukaryotic-type RPA homologue, the crenarchaeal SSB proteins appear to be much more related to the bacterial proteins, with a single OB fold and a flexible C-terminal tail. However, the crystal structure of the SSB protein from S. solfataricus showed that the OB-fold domain is more similar to that of the eukaryotic RPAs, supporting the close relationship between Archaea and Eukaryota. RPA from Methanosarcina acetivorans displays a unique property. Unlike the multiple RPA proteins found in other archaea and eukaryotes, each subunit of the M. acetivorans RPAs, RPA1, RPA2, and RPA3, have 4, 2, and 2 OB-folds, respectively, and can act as a distinct single-stranded DNA-binding proteins. Furthermore, each of the three RPA proteins, as well as their combinations, clearly stimulates the primer extension activity of M. acetivorans DNA polymerase BI in vitro , as shown previously for bacterial SSB and eukaryotic RPA. of SSB and RPA suggested that they are composed of different combinations of the OB fold. Bacterial and eukaryotic organisms contain one type of SSB or RPA, respectively. In contrast, archaeal organisms have various RPAs, composed of different organizations of OB-folds. A hypothesis that homologous recombination might play an important role in generating this diversity of OB-folds in archaeal cells was proposed, based on experiments characterizing the engineered RPAs with various OB-folds.
9. DNA polymerase
polymerase catalyzes phosphodiester bond formation between the terminal 3-OH of the primer and the?-Phosphate of the incoming triphosphate to extend...
