β-Crystallins of the adult chicken lens: Relatedness of the polypeptides and their aggregates

Abstract

β-Crystallins, the major class of polymeric soluble proteins in the mature lens, were isolated from one-year-old chickens by Sephadex G200 chromatography and identified by immunodiffusion. The β-crystallin polypeptides were resolved into six components with molecular weights near 35 K, 29 K, 26 K, 24 K, 20 K and 19 K by SDS-urea-polyacrylamide gel electrophoresis. The β-crystallins eluting later from the column contained relatively less of the 35 K polypeptides. Two-dimensional tryptic fingerprints of iodinated peptides indicated that the β-crystallin polypeptides had related primary structures, although some differences in the peptide maps were evident. Isoelectric focusing of undissociated β-crystallins in a sucrose density gradient resolved four major forms with isoelectric points of 4·6, 6·0, 6·4 and 6·6. Preparative isoelectric focusing proved to be an effective method for purification of these proteins. SDS-urea-polyacrylamide gel electrophoresis showed that each of the major isoelectric forms of β-crystallin had a characteristic combination of polypeptides. The 35 K and 29 K polypeptides were present only in the most acidic β-crystallins. Finally, circular dichroism spectroscopy showed that, despite their different polypeptide compositions, the four β-crystallins had similar secondary and tertiary conformations.