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L. J. Takemoto, K. Lampi, G. Wistow, L. Hancock; Differential Interaction of GammaS Crystallin With Wild Type and Mutant AlphaA Crystallin. Invest. Ophthalmol. Vis. Sci. 2007;48(13):2031. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
The transparent properties of the human lens are dependent upon short-range interactions of crystallins, which may involve noncovalent associations of alpha crystallins with gamma crystallins. Since a mutation of alphaA crystallin (R116C) causes cataract in humans, it is possible that the mutant protein may have altered interactions with gamma crystallins.
The wild type forms of human gammaS crystallin and alphaA crystallin, and the R116C mutant form of human alphaA crystallin, were expressed in E. coli, and then purified. Microequilibrium dialysis, followed by reverse phase chromatography, was used to determine possible noncovalent interactions of the alphaA crystallins with gammaS crystallin. Gel filtration chromatography under nondenaturing conditions, followed by reverse phase chromatography under denaturing conditions, was used to determine if the interactions of alphaA crystallins with gammaS crystallins were reversible.
Compared with wild type alphaA crystallin, microequilibrium dialysis showed that the R116C mutant of alphaA crystallin exhibited altered association with gammaS crystallin, under true equilibrium conditions. Purification of alphaA crystallin after equilibrium dialysis, followed by reverse phase chromatography, demonstrated that the alphaA-gammaS interactions were totally reversible.
Mutation of arginine to cysteine at residue 116 of human alphaA crystallin results in alteration of the interaction of alphaA crystallin with gammaS crystallin. This could contribute to changes in short-range order that eventually result in human cataractogenesis.
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