Abstract
Several mutations associated with congenital cataracts in human beings target conserved arginine residues in αA-crystallin. The N-terminal region of αA-crystallin is a "mutational hotspot," with multiple cataract-related mutations reported in this region. Two mutations at arginine 21 in the N-terminal domain of αA-crystallin - αA-R21L and αA-R21W have been associated with congenital cataract. A third mutant of R21, αA-R21Q, was recently identified to be associated with congenital cataract in a South Australian family. The point mutation was reported to compromise the quaternary structure of αA-crystallin by preventing its assembly into higher ordered oligomers. To assess the effect of the αA-R21Q mutation on αA-crystallin function, recombinant αA-R21Q was expressed, purified and characterized in vitro. Compared to wild-type αA-crystallin, the recombinant αA-R21Q exhibits enhanced chaperone-like activity, increased surface hydrophobicity, lesser stability in urea and increased susceptibility to digestion by trypsin. αA-R21Q demonstrated increased binding affinity towards unfolding ADH and bovine lens fiber cell membranes. αA-R21Q homo-oligomers and hetero-oligomers also prevented H(2)O(2)-induced apoptosis in ARPE-19 cells. Taken together, αA-R21Q exhibited a gain of function despite subtle structural differences as compared to wild-type αA-crystallin. This study further validates the involvement of arginine 21 in regulating αA-crystallin structure and function.