The alphaA-crystallin R116C mutant has a higher affinity for forming heteroaggregates with alphaB-crystallin.
An autosomal dominant congenital cataract in humans is associated with mutation of Arg-116 to Cys in alphaA-crystallin (alphaA-R116C). The chaperone activity and biophysical properties of reconstituted alpha-crystallin from different proportions of wild-type alphaB-crystallin (alphaB-wt) and alphaA-R116C-crystallin were studied by gel permeation chromatography, SDS-polyacrylamide gel electrophoresis, and fluorescence and circular dichroism spectroscopy and compared with those of reconstituted alpha-crystallin from alphaB-wt and wild-type alphaA-crystallin (alphaA-wt). The reconstituted alpha-crystallin containing alphaA-R116C and alphaB-wt had a higher molecular mass, a higher thermal sensitivity to exposition of Trp side chains, fewer available hydrophobic surfaces, and lower chaperone activity than the alpha-crystallin containing alphaA-wt and alphaB-wt. The secondary structure exhibited very small changes, whereas the tertiary structure was distinctly different for alpha-crystallin formed from alphaA-R116C and alphaB-wt. Most importantly, subunit exchange studies by fluorescence resonance energy transfer showed that alphaA-R116C forms heteroaggregates faster than alphaA-wt with alphaB-wt, and the reconstituted alpha-crystallins were true heteroaggregates of two interacting subunits. These findings suggest that the molecular basis for the congenital cataract with the alphaA-R116C mutation is the formation of highly oligomerized heteroaggregates of alpha-crystallin with modified structure. However, contrary to the earlier conclusions based on the studies of homoaggregates, the loss in chaperone activity of the heteroaggregates having alphaA-R116C does not appear to be large enough to become the main factor in initiating cataract development in the affected individuals.
Bera
Sibes
S
Department of Biochemistry and Molecular Biology, University of Arkansas for Medical Sciences, Little Rock, Arkansas 72205, USA.
Abraham
Edathara C
EC
D000817:Anilino Naphthalenesulfonates / Q000494:pharmacology; D000818:Animals; D001665:Binding Sites; D002942:Circular Dichroism; D003459:Crystallins / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000201:enzymology; D005456:Fluorescent Dyes / Q000494:pharmacology; D007700:Kinetics; D018832:Molecular Chaperones / Q000378:metabolism; D009154:Mutation; D011485:Protein Binding / Q000187:drug effects; D011487:Protein Conformation / Q000187:drug effects; D021122:Protein Subunits; D051381:Rats; D013696:Temperature; D013997:Time Factors
D000817:Anilino Naphthalenesulfonates; D003459:Crystallins; D005456:Fluorescent Dyes; D018832:Molecular Chaperones; D021122:Protein Subunits; C027132:1-anilino-8-naphthalenesulfonate