Resolution of Trp near UV CD spectra of calmodulin-domain peptide complexes into the 1La and 1Lb component spectra

Near uv Cd spectra of Trp residues in proteins frequently show a complex line shape deriving from the overlap of L-1(a) and L-1(b) electronic transitions. This study presents an original empirical method of resolving these components, based on the near uv CD spectra of well-defined complexes of calmodulin domains with target peptides containing a single Trp residue and derived from the skeletal muscle myosin light chain kinase target sequence. Spectra of 4 complexes were used to obtain the L-1(a) and L-1(b) component spectra that were then used to analyze further complexes. The broad and featureless L-1(a) spectrum is centered at 279 nm, the L-1(b) spectrum shows vibrational fine structure with maxima at 274.9, 281.5, and 289.8 nm. The CD spectrum of most complexes could successfully be fitted with one L-1(a) and one L-1(b) spectrum, the L-1(b) spectrum being negative for all complexes, however, failed to be adequately represented by only one L-1(a) and one L-1(b) spectrum. Instead, they could be fitted with one L-1(b) spectrum and two L-1(b) spectra with different sign and position. The method is successful in identifying and quantitating the relative intensities of a two-component system, consistent with a single conformation for tryptophan in a protein, and provides a simple indication of cases where a more complicated explanation is required. (C) 1998 John Wiley & Sons, Inc.