THE EFFECT OF CONCENTRATION ON THE STRUCTURE OF ALPHA-CRYSTALLIN

Several models have been proposed for arrangement of the subunits in a -crystallin. These include the contrasting proposals that subunits are arranged in three layers and that subunits assemble into micellelike structures. The validity of the micelle model was investigated by exam ining the effects of variations in protein concentration on the surfac e tension, conductivity, molecular weight and conformation of alpha-cr ystallin. The data were compared with those obtained for bovine serum albumin (BSA) and sodium dodecylsulphate (SDS). Measurements of surfac e tension were conducted in the range, 10 mu g ml(-1) to 130 mg ml(-1) , in low and high ionic strength buffers. An apparent point of inflect ion, independent of ionic strength, was seen in alpha-crystallin's sur face tension at around 1.9 mg ml(-1) (95 mu M). The surface tension di d not plateau beyond this point, as is the case with surfactants, but continued to decrease up to 130 mg ml(-1). BSA exhibited similar surfa ce tension properties with an apparent inflection at 0.9 mg ml(-1) (13 mu M). The conductivity of cc-crystallin and BSA solutions increased smoothly with no sign of any transition up to 96 mg ml(-1) and 60 mg m l(-1), respectively. In contrast, SDS showed a clear transition in thi s property at the concentration corresponding to its CMC. The aggregat ion state of the alpha-crystallin aggregates was examined by comparing molecular masses and Stokes radii. The size of the protein remained u niform over a wide concentration range and was unaffected by variation s in ionic strength, Protein conformation, which was monitored by exam ining the microenvironment of tryptophan residues, was also found to b e independent of protein concentration. It is concluded that over the concentration range that was investigated, alpha-crystallin does not e xhibit any of the properties associated with classical micelles formed from small amphiphilic molecules. (C) 1996 Academic Press Limited