The heteromeric BUF protein was originally shown to bind to URS1 elements which are situated upstream of many genes in Saccharomyces cerevisiae and mediate negative control of their transcription. Among the genes regulated through the URS1 site and the proteins interacting with it are those participating in carbon, nitrogen, and inositol metabolism; electron transport; meiosis; sporulation; and mating-type switching. We show here that pure BUF protein, in addition to binding to the negatively acting URS1 site, also binds to CAR1 sequences supporting transcriptional activation (upstream activation sequences). To determine the BUF protein structure, we cloned and sequenced the BUF1 and BUF2 genes and found them to be identical to the RF-A (RP-A) gene whose products participate in yeast DNA replication as single-stranded DNA binding proteins. These data argue that BUF protein-binding sites serve multiple roles in transcription and replication.