Direct NMR measurement of folding kinetics of a trimeric peptide.

Abstract

Direct NMR measurements of the folding kinetics are performed on a collagen-like triple helical peptide. The triple helical peptide was designed to model a biologically important region of collagen and has the sequence (POG)3ITGARGLAG(POG)4. Triple helical peptides were synthesized with specifically labeled 15N amino acid residues in key positions, and the kinetics of folding of the individual residues were monitored directly by measuring the loss of monomer intensity and the increase in trimer intensity. The residues at the terminal ends and central region could be followed independently and quantitated directly. Residues located at the terminal ends have rates and kinetics of folding that are distinct from residues in the central region of the peptide. This allows the monitoring of different steps in the folding mechanism and the postulation of the existence of a kinetic intermediate. The NMR data are consistent with a mechanism of association/nucleation and propagation. Hereditary connective tissue diseases are associated with mutations that result in abnormal folding of collagen, and the NMR folding experiments on a collagen-like peptide provide a basis for characterizing the molecular defect in folding mutations.