UC Irvine

Crystallins are structural proteins that serve as a medium for lens refraction to aid in the formation of vision. These highly soluble and stable proteins persist for decades without aggregation. Over the course of aging and exposure to degrading agents, such as reactive oxygen species, ultraviolet light, metal ions, or ionizing radiation, the eye lens crystallins accumulate post-translational modifications (PTMs) that can result in a loss of structural stability and thereby aggregation. These aggregates eventually form light-scattering masses and a diseased state of the eye lens known as cataract. Cataract is the leading cause of blindness in the world with millions affected every year. My research aims to evaluate the effects of PTMs on the structure and aggregation propensity of a specific human crystallin, γS-crystallin (γS). Two of the most common PTMs in aged and cataractous lenses are deamidation and oxidation. A series of deamidated variants of γS were studied via X-ray crystallography and biophysical characterization, revealing that the overall fold of γS is maintained among variants but these variants are structurally destabilized and more prone to disulfide bond formation. Additionally, the chemical modifications and stability of γS after exposure to ionizing radiation were evaluated. After high doses of γ radiation, γS accumulated a large number of modifications but largely resists unfolding. Both of these studies demonstrate the resilience of γS in spite of significant deamidation and oxidation modification, an adaptation advantageous for a long-lived protein. Lastly, a novel method of protein crystallization within the wells of a serial crystallography chip was developed. This technique lowers samples consumption and physical handling of potentially delicate crystals, expanding the scope of systems available to study via serial and time-resolved crystallography. A novel structure of a γS deamidation variant was solved via serial crystallography on microcrystals grown in-chip.