Abstract
Cataracts cause vision loss through the large-scale aggregation of eye lens proteins as a result of ageing or congenital mutations. The development of new treatments is hindered by uncertainty about the nature of the aggregates and their mechanism of formation. We describe the structure and morphology of aggregates formed by the P23T human γD-crystallin mutant associated with congenital cataracts. At physiological pH, the protein forms aggregates that look amorphous and disordered by electron microscopy, reminiscent of the reported formation of amorphous deposits by other crystallin mutants. Surprisingly, solid-state NMR reveals that these amorphous deposits have a high degree of structural homogeneity at the atomic level and that the aggregated protein retains a native-like conformation, with no evidence for large-scale misfolding. Non-physiological destabilizing conditions used in many in vitro aggregation studies are shown to yield qualitatively different, highly misfolded amyloid-like fibrils.
Highlights
Cataracts cause vision loss through the large-scale aggregation of eye lens proteins as a result of ageing or congenital mutations
At physiological pH, the 20.6 kDa P23T human gD-crystallin (hgD) is soluble at dilute concentrations, but undergoes a temperature-sensitive aggregation at 10–15 mg ml À 1 concentration (Supplementary Fig. 1)
We observed the aggregation of P23T hgD across a range of pH conditions, and found that the nature of the resulting aggregates is different at different pH values
Summary
Cataracts cause vision loss through the large-scale aggregation of eye lens proteins as a result of ageing or congenital mutations. Based on in vitro studies, two prominent mechanisms have been put forward to explain the aggregation propensity of mutated or modified crystallins associated with cataract formation. Reported arguments in favour of amyloid formation in vivo are largely based on the ability of mouse model cataracts to bind amyloid-binding dyes[31] Such dyes are not a foolproof measure of amyloid structure as both false positives and false negatives occur[33,34]. We study distinct aggregated states of the above-mentioned P23T hgD that is associated with congenital cataracts (Fig. 1) This mutant protein forms both worm-like fibrils and amorphous-looking aggregates, depending on the experimental conditions. These aggregates are shown by ssNMR to contain the aggregated protein in a well-ordered native-like conformation, with no evidence of dynamic or static disorder
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
