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. 1991 May 15;88(10):4089–4093. doi: 10.1073/pnas.88.10.4089

The multifunctional protein OBF1 is phosphorylated at serine and threonine residues in Saccharomyces cerevisiae.

S C Francesconi 1, S Eisenberg 1
PMCID: PMC51603  PMID: 2034654

Abstract

We have purified a DNA replication enhancer-binding protein, OBF1, from yeast cells grown in a medium containing 32P-labeled orthophosphate. The purified 32P-labeled protein comigrated on polyacrylamide gels with OBF1 bands identified by immunoblotting with anti-OBF1 antibodies. Furthermore, trypsin treatment of the 32P-labeled OBF1 revealed several phosphorylated peptides, suggesting that OBF1 is multiply phosphorylated in vivo. Incubation of phosphorylated peptides with calf intestinal phosphatase liberated the radiolabel as free phosphate, indicating a phosphoester linkage. Acid hydrolysis of the tryptic peptides revealed 32P-label label comigrating with phosphoserine; some of it, however, was also identified as phosphothreonine. Using anti-OBF1 antibodies, we cloned the OBF1 gene from a lambda gt11 yeast expression library. The DNA sequence of the isolated gene and its over-expression in yeast indicated that OBF1 is identical to ABF-1 and BAF1 proteins, believed to have a role in transcriptional repression and activation. Therefore, we suggest that OBF1 is a multifunctional protein, acting in transcription and replication, and that these activities are regulated by phosphorylation.

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