Spectral contribution of the individual tryptophan of alpha B-crystallin: A study by site-directed mutagenesis

There are two tryptophan residues in the lens alpha B-crystallin, Trp9 and Trp60. We prepared two Trp --> Phe substituted mutants, W9F and W60F, for u se in a spectroscopic study. The two tryptophan residues contribute to Trp fluorescence and near-ultraviolet circular dichroism (UV CD) differently. T he major difference in the near-UV CD is the contribution of L-1(a) of Trp: it is positive in W60F but becomes negative in W9E Further analysis of the near-UV CD shows an increased intensity in the region of 270-280 nm for W6 0F, suggesting that the Tyr48 is affected by the W60F mutation. It appears that Trp60 is located in a more rigid environment than Trp9, which agrees w ith a recent structural model in which Trp60 is in a beta-strand.