Properties of single-stranded DNA-binding proteins (SSB-proteins) from chromatin and nonchromatin fractions of Ehrlich ascites tumour: phosphorylation enhances the affinity of SSB-proteins for single-stranded DNA.

To assess the possible functional role of single-strand DNA-binding (SSB) proteins in eucaryotic cell, a comparative study was made of SSB-proteins isolated from chromatin and the nonchromatin fractions of Ehrlich ascites tumour (EAT) cells. No appreciable differences between the two groups could be found either in SDS-gel electrophoretic patterns or in the ssDNA-binding capacity and stimulation of DNA replication in permeable EAT cells. However, the chromatin SSB-proteins incorporated 1.4-times more labelled phosphate in vivo; phosphate assays in the isolated chromatin and nonchromatin SSB-proteins yielded ca. 3 and 2 moles Pi/mole protein, respectively. Both preparations could be further phosphorylated in vitro with Ca-phospholipid-dependent protein kinase and the catalytic subunit of cAMP-dependent protein kinase, but the non-chromatin proteins were phosphorylated to a greater degree. In parallel with phosphorylation, the SSB-proteins displayed stronger binding to ssDNA cellulose. Phosphorylation may thus be a means of regulating the functions of SSB-proteins, in particular their interaction with chromatin.

Relationship of single-stranded DNA-binding proteins of Ehrlich ascites tumour to cell growth phase and DNA replications.

The possible involvement of SSB-proteins in DNA replication in Ehrlich ascites tumour (EAT) has been investigated. A direct relation (the computer-generated correlation coefficient was 0.9) between the SSB-proteins content in chromatin and intensity of the replicative synthesis of DNA in various preparation of EAT in vivo and in vitro is observed. Addition of exogenous SSB-proteins to the permeable EAT cells has been found to increase the replicative synthesis. Although eukaryotic SSB-proteins are not complete analogs of the prokaryotic SSB-proteins, they evidently participate in DNA replication in eukaryotic cells and possibly are intracellular regulators of proliferation.