Thermodynamic stability of human lens recombinant alpha A- and alpha B-crystallins

Lens alpha-crystallin is a 600-800-kDa heterogeneous oligomer protein consi sting of two subunits, alpha A and alpha B. The homogeneous oligomers (alph a A- and alpha B-crystallins) have been prepared by recombinant DNA technol ogy and shown to differ in the following biophysical/biochemical properties : hydrophobicity, chaperone-like activity, subunit exchange rate, and therm al stability. In this study, we studied their thermodynamic stability by un folding in guanidine hydrochloride. The unfolding was probed by three spect roscopic parameters: absorbance at 235 nm, Trp fluorescence intensity at 32 0 nm, and far-UV circular dichroism at 223 nm, Global analysis indicated th at a three-state model better describes the unfolding behavior than a two-s tate model, an indication that there are stable intermediates for both alph a A- and alpha B-crystallins, In terms of standard free energy (Delta G(NU) (H2O)), alpha A-crystallin is slightly more stable than alpha B-crystallin, The significance of the intermediates may be related to the functioning of cu-crystallins as chaperone-like molecules.