Structure and activity associated with multiple forms of Schizosaccharomyces pombe DNA polymerase δ Journal Article
| Authors: | Zuo, S.; Bermudez, V.; Zhang, G.; Kelman, Z.; Hurwitz, J. |
| Article Title: | Structure and activity associated with multiple forms of Schizosaccharomyces pombe DNA polymerase δ |
| Abstract: | DNA polymerase δ (Pol δ) isolated from Schizosaccharomyces pombe (sp) consists of at least four subunits, Pol3, Cdc1, Cdc27, and Cdm1. We have reconstituted the four-subunit complex by simultaneously expressing these polypeptides in baculovirus-infected insect cells. The properties of the purified cloned spPol δ were identical to the native spPol δ isolated from S. pombe cells. In addition, we also isolated a three-subunit complex containing Pol3, Cdc1, and Cdm1. Both three- and four-subunit complexes required replication factor C and proliferating cell nuclear antigen for DNA replication. However, in the presence of low levels of polymerase complexes, the three-subunit complex was less efficient than the four-subunit complex in supporting DNA replication. The inefficient synthesis of DNA by the three- subunit complex can be remedied by the addition of Cdc27, the subunit missing in the three-subunit complex. Gel filtration analysis demonstrated that the three-subunit complex is a monomer of the heterotrimer (Pol3, Cdc1, and Cdm1) and that the four-subunit complex is a dimer of the heterotetramer (Pol3, Cdc1, Cdc27, and Cdm1), similar to the structure of native spPol δ. We have further shown that Cdc1 and Cdc27 interact to form a heterodimeric complex. Gel filtration studies indicate that the structure of this complex is dimeric. These observations suggest that the Cdc27 subunit may play an important role contributing to the dimerization of Pol δ. |
| Keywords: | controlled study; protein expression; dna replication; dna synthesis; cell cycle proteins; enzyme activity; structure-activity relationship; dna; protein purification; recombinant proteins; dimerization; replication factor c; cycline; structure analysis; enzyme structure; enzyme subunit; isoenzymes; schizosaccharomyces; schizosaccharomyces pombe proteins; protein isolation; schizosaccharomyces pombe; dna directed dna polymerase delta; dna polymerase iii; unidentified baculovirus; insecta; priority journal; article |
| Journal Title: | Journal of Biological Chemistry |
| Volume: | 275 |
| Issue: | 7 |
| ISSN: | 0021-9258 |
| Publisher: | American Society for Biochemistry and Molecular Biology |
| Date Published: | 2000-02-18 |
| Start Page: | 5153 |
| End Page: | 5162 |
| Language: | English |
| DOI: | 10.1074/jbc.275.7.5153 |
| PUBMED: | 10671561 |
| PROVIDER: | scopus |
| DOI/URL: | |
| Notes: | Export Date: 18 November 2015 -- Source: Scopus |
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