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 1La and 1Lb 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 1La and 1Lb component spectra that were then used to analyze further complexes. The broad and featureless 1La spectrum is centered at 279 nm, the 1Lb 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 1La and one 1Lb spectrum, the 1Lb spectrum being negative for all complexes but the 1La spectrum showing either positive or negative sign. Spectra of some complexes, however, failed to be adequately represented by only one 1La and one 1Lb spectrum. Instead, they could be fitted with one 1Lb spectrum and two 1La 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.
Barth
A
A
Division of Physical Biochemistry, National Institute for Medical Research, London, UK.
Martin
S R
SR
Bayley
P M
PM
D000818:Animals; D002147:Calmodulin / Q000737:chemistry*; D002942:Circular Dichroism; D004331:Drosophila melanogaster / Q000737:chemistry; D004926:Escherichia coli / Q000235:genetics; D008968:Molecular Conformation; D018482:Muscle, Skeletal / Q000737:chemistry; D009219:Myosin-Light-Chain Kinase / Q000378:metabolism; D010446:Peptide Fragments / Q000737:chemistry*; D011994:Recombinant Proteins / Q000737:chemistry; D014364:Tryptophan / Q000737:chemistry*
D002147:Calmodulin; D010446:Peptide Fragments; D011994:Recombinant Proteins; D014364:Tryptophan; D009219:Myosin-Light-Chain Kinase