C-terminal lysine truncation increases thermostability and enhances chaperone-like function of porcine αB-crystallin

Abstract

The carboxyl-terminal segment of α-crystallin, a major lens protein of all vertebrates, has a short and flexible peptide extension of about 20 amino acid residues that are very susceptible to proteolytic truncation and modifications under physiological conditions. To investigate its role in crystallin aggregation and chaperone-like activity, we constructed a mutant of porcine αB-crystallin with C-terminal lysine truncated end, which unexpectedly showed better chaperone-like function than wild-type αB-crystallin. From circular dichroism (CD) spectra, we show that the mutant possesses similar secondary and tertiary structures to those of native purified and recombinant αB-crystallins. Analytical ultracentrifugation revealed that the truncated mutant was smaller than wild-type αB-crystallin in aggregation size and mass. The observed higher thermostability and anti-thermal aggregation propensity of the truncated αB-crystallin mutant than wild-type αB-crystallin are in contrast to the prevailing notion that mutations at the C-terminal lysines of αB-crystallin result in substantial loss of chaperone-like activity, despite the overall preservation of secondary structure. The detailed characterization of the C-terminal deletion mutants may provide some deeper insight into the chaperoning mechanism of the structurally related small heat-shock protein family.