Effect of Structural Changes Induced by Deletion of 54FLRAPSWF61 Sequence in αB-crystallin on Chaperone Function and Anti-Apoptotic Activity

Previously, we showed that the removal of the 54-61 residues from alpha B-crystallin (alpha B triangle 54-61) results in a fifty percent reduction in the oligomeric mass and a ten-fold increase in chaperone-like activity. In this study, we investigated the oligomeric organization changes in the deletion mutant contributing to the increased chaperone activity and evaluated the cytoprotection properties of the mutant protein using ARPE-19 cells. Trypsin digestion studies revealed that additional tryptic cleavage sites become susceptible in the deletion mutant than in the wild-type protein, suggesting a different subunit organization in the oligomer of the mutant protein. Static and dynamic light scattering analyses of chaperone-substrate complexes showed that the deletion mutant has more significant interaction with the substrates than wild-type protein, resulting in increased binding of the unfolding proteins. Cytotoxicity studies carried out with ARPE-19 cells showed an enhancement in anti-apoptotic activity in alpha B triangle 54-61 as compared with the wild-type protein. The improved anti-apoptotic activity of the mutant is also supported by reduced caspase activation and normalization of the apoptotic cascade components level in cells treated with the deletion mutant. Our study suggests that altered oligomeric assembly with increased substrate affinity could be the basis for the enhanced chaperone function of the alpha B triangle 54-61 protein.

Automated summary

The deletion of residues 54-61 from alpha B-crystallin results in increased chaperone activity and altered oligomeric assembly, with a different subunit organization observed in the mutant protein. The mutant protein shows more significant interaction with substrates and increased binding of the unfolding proteins compared to the wild-type protein. Additionally, the mutant exhibits enhanced anti-apoptotic activity, supported by reduced caspase activation and normalization of apoptotic cascade components in cells.