Refinement of 3D structure of bovine lens αA-crystallin

Abstract

In absence of 3D structures for α-crystallin subunits, αA and αB, we utilized a number of experimental and molecular modeling techniques to generate working 3D models of these polypeptides (Farnsworth et al., 1994. In Molecular Modeling: From Virtual Tools to Real Problems (Eds. Kumosinski, T.F. and Liebman, M.N.) ACS Symposium Series 576, Ch. 9:123–134, 1994, ACS Books, Washington DC). The refinement of the initial bovine αA 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 predictions in combination with database searches. The spectroscopic study reveals that α-crystallin is not an all β-sheet protein but contains ∼17% α-helices, ∼33% β-structures and ∼50% turns and coils. The refinement of the αA structure results in an elongate, asymmetric amphipathic molecule. The hydrophobic N-terminal domain imparts the driving force for subunit aggregation 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 αA, the highly negatively charged C-terminal domain has three small positive areas which may participate in dimer or tetramer formation of independently expressed C-terminal domains. The electrostatic potential of positive areas is modulated and become more negative with phosphorylation and ATP binding. The refined bovine αA model was used to construct αA models for the human, chick and dogfish shark. A high degree of conservation of the three dimensional structure and the electrostatic potential was observed. Our proposed open micellar quaternary structure correlates well with experimental data accumulated over the past several decades. The structure is also predictive of the more recent data.