Limited proteolysis of γII‐crystallin from calf eye lens

摘要: γII-crystallin from calf eye lens consists of two homologous domains, connected by a six-residue linker peptide. In order to study the intrinsic properties domains and their mutual stabilization, limited proteolysis was applied. Optimum conditions providing homogeneous 10-kDa fragment at high yield were pepsin cleavage in 0.1 M NaCl/HCl pH 2.0, presence 3.0 urea. Determination N-terminus C-terminal sequence showed that occurred Phe88–Arg89 peptide bond, giving rise complete N-terminal domain including connecting hexapeptide. The part polypeptide chain is cleaved small fragments. Comparing spectral isolated intact proved structure be closely similar native domain. Small differences absorbance, fluorescence emission circular dichroism point alterations caused increase surface area as consequence separation. resistance toward thermal alkaline denaturation, well unfolding urea or guanidine · HCl decreased, due lack interactions stabilizing protein. Unfolding/folding kinetics coincide with second phase bimodal transition γII-crystallin, agreement independent sequential folding modular assembly within molecule.