Purification and characterization of a nuclear DNA-binding phosphoprotein in fetal and tumor tissues.
The basic nonhistone phosphoprotein 110/8.4 (M.W. X 10(-3)/pI) was found in 0.35 M NaCl nuclear extracts of four tumor tissues, i.e., fast-growing Novikoff hepatoma, Morris hepatoma 3924A, HeLa cells, and Namalwa cells; it was also found in fetal rat liver. This protein was not detected in normal or regenerating liver and thus may represent an "oncofetal" protein of potential interest as a cancer "marker." Protein 110/8.4 was purified approximately 4000- to 5000-fold under nondenaturing condition from 0.35 M NaCl nuclear extracts of Novikoff hepatoma cells or Namalwa cells by ammonium sulfate fractionation, calcium phosphate gel treatment, and phosphocellulose chromatography. Sodium dodecyl sulfate:polyacrylamide gel electrophoresis of the purified native protein revealed a single polypeptide chain with a molecular weight of approximately 110,000. The pI of the protein was estimated to be 8.4 by nonequilibrium pH gradient electrophoresis in 9 M urea; accordingly, this protein was designated 110/8.4. Amino acid analysis showed that Protein 110/8.4 had an acidic:basic amino acid ratio of 1.25 and a high lysine and serine content; approximately 20% of the serine residues were found to be phosphorylated. Hydrazinolysis indicated that the carboxyl-terminal amino acid was serine; the amino terminus appeared to be blocked. Binding of Protein 110/8.4 to DNA was studied by the nitrocellulose filter assay. High-affinity binding occurred at ionic strength equal to or below 0.15 M.
D000818:Animals; D002467:Cell Nucleus / Q000032:analysis; D002868:Chromosomal Proteins, Non-Histone / Q000302:isolation & purification*; D004273:DNA, Neoplasm / Q000378:metabolism; D007526:Isoelectric Point; D008297:Male; D008970:Molecular Weight; D009363:Neoplasm Proteins / Q000302:isolation & purification*; D009374:Neoplasms, Experimental / Q000032:analysis* / Q000648:ultrastructure; D009994:Osmolar Concentration; D010750:Phosphoproteins / Q000302:isolation & purification*; D051381:Rats; D014018:Tissue Distribution
D002868:Chromosomal Proteins, Non-Histone; D004273:DNA, Neoplasm; D009363:Neoplasm Proteins; D010750:Phosphoproteins