Purification and partial characterization of a 19 KD/pI 4.5 nucleolar phosphoprotein

Abstract

Two-dimensional PAGE analysis of proteins associated with the slowly sedimenting "fibrillar" structures of HeLa nucleoli revealed a protein with a M_r of 19,000 and a pI of 4.5 which was highly labeled both with ³²P-orthophosphate and ³⁵S-methionine. The protein was isolated from Novikoff hepatoma nucleoli by extraction in 0.35 M NaCl and 5 mM DTT followed by chromatography in EDTA on DEAE-cellulose and Sephadex G-100. The protein was homogeneous with respect to two-dimensional PAGE, number of tryptic peptides and carboxyl terminal analysis. The protein contained an acidic/basic amino acid ratio of 2.1, 7 residues of methionine, 2 residues of cysteine, a blocked amino terminus and a carboxyl terminal lysylleucine.