Effects of molecular association on structure and dynamics of a collagenous peptide

Peptide GVKGDKGNPGWPGAPY from the triple-helix domain of type IV collagen aggregates in solution at a critical aggregation concentration of 18 mM. This molecular self-association process is investigated by 1H- and 13C-nmr spectroscopy. As a function of increasing peptide concentration, selective 1H resonances are cooperatively chemically shifted by up to 0.04 ppm to apparently saturable values at high concentration. Pulsed field gradient nmr was used to derive translation diffusion constants that, as the peptide concentration is increased, also cooperatively and monotonically decrease to an apparent limiting value. An average number of 6 monomer units per aggregate have been estimated from diffusion constant and 13C relaxation data. Comparative 1H nuclear Overhauser effect spectroscopy (NOESY) spectra accumulated at high and low peptide concentrations suggest that average internuclear distances are decreased as a result of peptide association. 13C-nmr multiple spin-lattice relaxation and 13C-[1H] NOE effects on 13C-enriched glycine methylene positions in the peptide demonstrate that overall molecular tumbling and backbone internal motions are attenuated in the aggregate state. Lowering the solution pD from pD 6 to pD 2 disrupts the aggregate state, suggesting a role for electrostatic interactions in the association process. Based on thermodynamic considerations, hydrophobic interactions also probably act to stabilize the aggregate state. These data are discussed in terms of an nmr/NOE constrained computer-modeled structure of the peptide.