COMPARISON OF THE 10000 AND 43000 DALTON POLYPEPTIDE POPULATIONS ISOLATED FROM THE WATER-SOLUBLE AND INSOLUBLE FRACTIONS OF HUMAN CATARACTOUS LENSES

The 10 000 and 43 000 dalton polypeptides from pooled human cataractous lenses were isolated from the water soluble fraction and were compared with the respective polypeptides isolated from the water insoluble fraction. The water soluble proteins were first separated with Sephadex G-200 under non-deaggregating conditions. The 10 000 dalton polypeptide was found principally in the last major protein component eluting from the column. The 43 000 dalton polypeptide was found apparently complexed non-covalently primarily within three aggregate populations ranging in size from 4 to 15 × 104 daltons. The 43 000 as well as the 10 000 dalton polypeptides from the G-200 column were purified in acetic acid-urea on Sephadex G-100 and G-150, respectively. The 10 000 dalton polypeptide can also be purified after the initial column fractionation without dissociation. The polypeptides from the water insoluble fraction were isolated by methodologies previously reported. Characterization of the 10 000 and 43 000 dalton polypeptides isolated from the water soluble and insoluble fractions indicated marked similarities. However, with both polypeptide populations the water insoluble component contained a greater negative charge, lower abundance of certain amino acids (e.g. tryptophan, tyrosine, methionine), and a greater atypical fluorescence. Immunochemical data indicates that the 10 000 dalton population from both soluble and insoluble fractions contains components arising from a number of different polypeptides including soluble crystallins and the extrinsic 43 000 dalton polypeptide. It was found that there is complete immunochemical identity between the soluble 43 000 dalton polypeptide and its counterpart in the disulfide high molecular weight species and the species isolated from the urea extracted nonreduced water insoluble fraction. However, the urea extracted reduced water insoluble fraction reacted only to the extent of 30% with the anti-water soluble 43 000 dalton polypeptide suggesting at least one other 43 000 dalton component which is present only in the water insoluble fraction. © 1979.