Heteromeric formation with βA3 protects the low thermal stability of βB1-L116P

Background/aims Congenital cataract is the leading cause of visual disability and blindness in childhood. beta B1-crystallin (CRYBB1) comprises about 1/10th of crystallin structural proteins, forming heteromers to maintain lens transparency. We previously reported a CRYBB1 mutation (c.347T>C, p.L116P) affecting 16 patients in a congenital nuclear cataract family. In this study, we investigate the underlying pathogenic mechanism of beta B1-L116P. Methods Protein isolation, size-exclusion chromatography, spectroscopy, Uncle stability screens and molecular dynamics simulations were used to assess beta A3- and beta B1-crystallin thermal stability, structural properties and heteromer formation. Results Cells that overexpressed beta B1-L116P tended to form aggregates and precipitations under heat-shock stress. Thermal denaturation and time-dependent turbidity experiments showed that thermal stability was significantly impaired. Moreover, protein instability appeared to increase with elevated concentrations detected by the Uncle system. Additionally, beta A3 had a relative protective effect on beta B1-L116P after heteromers were formed, although beta A3 was relatively unstable and was usually protected by basic beta-crystallins. Molecular dynamic simulations revealed that L116P mutation altered the hydrophobic residues at the surface around the mutant site, providing solvents more access to the internal and hydrophobic parts of the protein. Conclusions Decreased beta B1-crystallin thermal stability in the presence of the cataract-related L116P mutation contributes significantly to congenital cataract formation. Moreover, its formation of heteromers with beta A3 protects against the low thermal stability of beta B1-L116P.

Automated summary

This study reveals that the decreased thermal stability of beta B1-crystallin due to the L116P mutation contributes significantly to the formation of congenital cataracts. Additionally, the heteromer formation between beta B1 and beta A3 protects against the low thermal stability of beta B1-L116P.