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| A hierarchy of SSB protomers in replication protein A |
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| Author(s): Philipova D, Mullen JR, Maniar HS, Lu JA, Gu CY, Brill SJ |
| Source: GENES & DEVELOPMENT Volume: 10 Issue: 17 Pages: 2222-2233 Published: SEP 1 1996 |
| Times Cited: 100 References: 56 |
| Abstract: Replication Protein A (RPA) is a heterotrimeric single-stranded DNA-binding protein (SSB) found in ail eukaryotic cells. RPA is known to be required for many of the same reactions catalyzed by the homotetrameric SSB of bacteria, but its origin, subunit functions, and mechanism of binding remain a mystery. Here we show that the three subunits of yeast RPA contain a total of four domains with weak sequence similarity to the Escherichia coli SSB protomer. We refer to these four regions as potential ssDNA-binding domains (SBDs). The p69 subunit, which is known to bind ssDNA on its own, contains two SBDs that together confer stable binding to ssDNA. The p36 and p13 subunits each contain a single SBD that does not bind stably, but corresponds to the minimal region required for viability in yeast. Photocross-linking of recombinant protein to ssDNA indicates that an SBD consists of similar to 120 amino acids with two centrally located aromatic residues. Mutation of these aromatic residues inactivates ssDNA binding and is a lethal event in three of the four domains. Finally, we present evidence that the p36 subunit binds ssDNA, as part of the RPA complex, in a salt-dependent reaction similar to the wrapping of ssDNA about E. coli SSB. The results are consistent with the notion that RPA arose by duplication of an ancestral SSB gene and that tetrameric ssDNA-binding domains and higher order binding are essential features of cellular SSBs. |
| Document Type: Article |
| Language: English |
Addresses:
1. RUTGERS STATE UNIV, DEPT MOL BIOL & BIOCHEM, CTR ADV BIOTECHNOL & MED, PISCATAWAY, NJ 08855 USA |
| Publisher: COLD SPRING HARBOR LAB PRESS, 1 BUNGTOWN RD, PLAINVIEW, NY 11724 |
| Subject Category: Cell Biology; Developmental Biology; Genetics & Heredity |
| IDS Number: VG107 |
| ISSN: 0890-9369 |
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