Characterization of βB2‐crystallin tryptophan mutants reveals two different folding states in solution

Abstract

AbstractConserved tryptophan residues are critical for the structure and the stability of β/γ‐crystallin in the lenses of vertebrates. During aging, in which the lenses are continuously exposed to ultraviolet irradiation and other environmental stresses, oxidation of tryptophan residues in β/γ‐crystallin is triggered and impacts the lens proteins to varying degrees. Kynurenine derivatives, formed by oxidation of tryptophan, accumulate, resulting in destabilization and insolubilization of β/γ‐crystallin, which correlates with age‐related cataract formation. To understand the contribution of tryptophan modification on the structure and stability of human βB2‐crystallin, five tryptophan residues were mutated to phenylalanine considering its similarity in structure and hydrophilicity to kynurenine. Among all mutants, W59F and W151F altered the stability and homo‐oligomerization of βB2‐crystallin—W59F promoted tetramerization whereas W151F blocked oligomerization. Most W59F dimers transformed into tetramer in a month, and the separated dimer and tetramer of W59F demonstrated different structures and hydrophobicity, implying that the biochemical properties of βB2‐crystallin vary over time. By using SAXS, we found that the dimer of βB2‐crystallin in solution resembled the lattice βB1‐crystallin dimer (face‐en‐face), whereas the tetramer of βB2‐crystallin in solution resembled its lattice tetramer (domain‐swapped). Our results suggest that homo‐oligomerization of βB2‐crystallin includes potential inter‐subunit reactions, such as dissociation, unfolding, and re‐formation of the dimers into a tetramer in solution. The W>F mutants are useful in studying different folding states of βB2‐crystallin in lens.