Fig. 1. Chromatography of proteolytic hydrolysates of lamins B and C on Sephadex LH-20.

HeLa cells were labeled with [³H]MVA or [³⁵S]cysteine, lamins B and C were isolated, solubilized in SDS, and extensively hydrolyzed with a combination of proteases, and the hydrolysates were concentrated on DEAE-Sephacel. 65% of the radioactivity in the hydrolysate of ³H-labeled lamin B adsorbed onto DEAE and could subsequently be eluted with formic acid/ethanol/water (2:8:1). In contrast, only 13 and 11%, respectively, of the radioactivity from the hydrolysates of ³⁵S-labeled lamin B and lamin C behaved similarly. The figure shows chromatograms that were obtained when the DEAE-eluted, ³H- and ³⁵S-labeled materials were each passed separately through Sephadex LH-20 in 20% formic acid in ethanol. Panel A, filled circles, ³H-labeled material from hydrolysate of lamin B (145,000 cpm loaded); open circles, corresponding, ³⁵S-labeled material (100,000 cpm loaded). Panel B, ³⁵S-labeled material from lamin C (80,000 cpm loaded). Note that an authentic standard of S-farnesyl cysteine coeluted with the major peak of ³H- label, shown in panel A, while cysteine and cystine emerged in the position indicated by the arrow. Hydrolysates of labeled lamin B yielded results like these in three different experiments, although the small peak of ³H-labeled material, that emerged at an elution volume of 20 ml, contained variable proportions (5–20%) of the total radioactivity. Hydrolysates of the [³H]MVA-labeled, minor nuclear protein of 66 kDa, described previously (5) also yielded results that were similar to those shown in panel A.