Comparative analysis of the amino- and carboxy-terminal domains of calmodulin by Fourier transform infrared spectroscopy.

Fourier transform infrared spectra were obtained for mammalian calmodulin and two of its fragments produced by limited proteolysis with trypsin TR1C (1-77) and TR2C (78-148). Experiments were done in H2O, D2O and D2O/trifluoroethanol (TFE) mixtures. Information about secondary structure was obtained from analysis of the amide I and II bands; while characteristic absorbances for tyrosine, phenylalanine and carboxylate groups were analyzed for changes in tertiary structure. Our data indicate that the secondary and tertiary structure is preserved in the two half molecules of CaM, both in the apo- and Ca(2+)-saturated state. Addition of the structure-inducing solvent TFE causes marked changes only in the apo-TR1C domain. The maximum wavenumber for the amide I band of the two domains of CaM in D2O was markedly different (1642 cm-1 for TR1C versus 1646/1648 cm-1 for Ca2+ and apo-TR2C). This renders the amide I band for the intact protein very broad in comparison to that in other proteins and is indicative of a distribution of alpha-helices with slightly different hydrogen bonding patterns.