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Mutations associated with early cataract development in mice destabilize human gammaD‐crystallin
Author(s) -
Drahos Kate Lauren,
King Jonathan
Publication year - 2008
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.22.2_supplement.232
Subject(s) - crystallin , mutant , chemistry , mutation , transition (genetics) , protein aggregation , protein folding , biophysics , microbiology and biotechnology , biology , biochemistry , gene
Three mutations, identified by genetic screens for cataract in mice, were selected and homologous mutations were made in the human lens protein γD‐crystallin (HγD). HγD is a monomeric, two‐domain protein found in the nuclear region of the lens. The three mutant proteins produced were HγD L5S, V75D, and I90F. Equilibrium unfolding/refolding experiments were performed to measure the stability of the mutant proteins compared to wildtype (wt) HγD. wt‐HγD has been previously shown to exhibit a three‐state unfolding/refolding transition. Two of the mutant proteins, L5S and V75D, also show three‐state unfolding/refolding transitions with significantly populated intermediates. In both cases, the first transition midpoint is shifted to lower denaturant concentrations. The third mutant protein, I90F, exhibits a two‐state equilibrium unfolding/refolding transition. Like wt‐HγD, all three mutants appear to aggregate upon refolding. However, unlike wt‐HγD, L5S and V75D do not appear to refold to a native‐like state before aggregating. All three mutations destabilize the protein compared to wt‐HγD and aggregate upon refolding, consistent with the phenotype observed in mice of early cataract development. These studies will help to further our understanding of how, at the molecular level, these mutations result in the eye disease cataract. This research was supported by NIH (GM17980) and NEI (EY015834) to J.K.