REFINEMENT OF 3D STRUCTURE OF BOVINE LENS ALPHA-A-CRYSTALLIN

In absence of 3D structures for alpha-crystallin subunits, alpha A and alpha B, we utilized a number of experimental and molecular modeling techniques to generate working 3D models of these polypeptides (Farnsw orth et al., 1994. In Molecular Modeling: From Virtual Tools to Real P roblems (Eds. Kumosinski, T.F. and Liebman, M.N.) ACS Symposium Series 576, Ch. 9:123-134, 1994, ACS Books, Washington DC). The refinement o f the initial bovine alpha A model was achieved using a more accurate estimation of secondary structure by new/updated methods for analyzing the far UV-CD spectra and by neural network secondary structure predi ctions in combination with database searches. The spectroscopic study reveals that alpha-crystallin is not an all beta-sheet protein but con tains similar to 17% alpha-helices, similar to 33% beta-structures and similar to 50% turns and coils. The refinement of the alpha A structu re results in an elongate, asymmetric amphipathic molecule. The hydrop hobic N-terminal domain imparts the driving force for subunit aggregat ion while the more flexible, polar C-terminal domain imparts aggregate solubility. In our quaternary structure of the aggregate, the monomer is the minimal cooperative subunit. In bovine alpha A, the highly neg atively charged C-terminal domain has three small positive areas which may participate in dimer or tetramer formation of independently expre ssed C-terminal domains. The electrostatic potential of positive areas is modulated and become more negative with phosphorylation and ATP bi nding. The refined bovine alpha A model was used to construct alpha A models for the human, chick and dogfish shark. A high degree of conser vation of the three dimensional structure and the electrostatic potent ial was observed. Our proposed open micellar quaternary structure corr elates well with experimental data accumulated over the past several d ecades. The structure is also predictive of the more recent data. (C) 1998 Elsevier Science B.V. All rights reserved.