Isolation and partial characterization of a nuclear antigen (68/6.3) from the Namalwa cell line (a Burkitt lymphoma).

Abstract

A nuclear antigen was purified from the 0.01 M Tris-HCl/pH8 extract of nuclei of the Burkitt tumor Namalwa cell line to electrophoretic homogeneity by DEAE cellulose chromatography, affinity chromatography, and preparative isoelectric focusing. The yield of antigens was 0.02% of the nuclear 0.01 M Tris-HCl/pH8 extract. On two-dimensional gel electrophoresis, the major antigen separated into two adjacent protein spots with molecular weights of 68,000 and an approximate pI of 6.3 (68/6.3 A and 68/6.3 B). A minor antigen had a molecular weight of 61,000 and pI of 6.0 (61/6.0). Fourteen 125I-labeled peptides were obtained from the tryptic digest of the major antigen (68/6.3 A and 68/6.3 B). The amino-acid composition analysis of the purified antigens indicated that the amino acids in the highest content were glycine (15%), glutamic acid (11.6%), and serine (9%); the ratio of acidic to basic amino acids was 1.95. In studies on nucleolytic activity, the purified antigen produced a single-stranded and then a double-stranded cleavage of PM 2 and pBR 322 DNA. This antigen is the first purified nuclear antigen that reacts with the HeLa-specific nucleolar antibodies.