The structure of two alanine containing ferrichromes: sequence determination by proton magnetic resonance.

Metal coordination confers an extraordinary structural stability to the ferrichromes which, independent of their variable amino acid composition, results in a basically unperturbed conformation for all the homologous peptides in the series. The proton magnetic resonance (pmr) characteristics for Al3+ analogues (alumichromes) reflect this conformational isomorphism in usual solvents so that single site substitutions are clearly recognized in the pmr spectra. Thus, the substitution of glycine by L-alanine or L-serine introduce new resonances characteristic of the sidechains and alter the pattern of the amide NH pmr region in that doublets substitute for glycyl triplets at the same site. Since for glycine- and L-serine containing alumichromes the resonances have already been identified, it is possible to unequivocally establish the primary structure of the two L-alanyl homologues ferrichrome C (see article) and sake colorant A (see article) on the basis of the comparative pmr spectra of their Al3+ analogues, namely, alumichrome C and alumisake. The resonance assignment, and hence the site occupancy, is substantiated by the temperature coefficients of the NH chemical shifts, rates of 1H-2H exchange and homonuclear proton spin decoupling experiments centered on the NH spectral region. Occupancy of site 1 by a glycine residue is observed for all known ferrichromes, which serves to conserve a "hairpin" turn. This method of obtaining sequence information should prove of general use for other systems of homologous polypeptides, provided their conformations are not affected by the residue substitutions.