13C-NMR relaxation study of heparin-disaccharide interactions with tripeptides GRG and GKG

Dmitri Mikhailov, Kevin H Mayo, Azra Pervin, Robert J. Linhardt

Research output: Contribution to journalArticlepeer-review

18 Scopus citations

Abstract

Heparin is a polydisperse sulphated copolymer consisting mostly of 1 → 4 linked glucosamine and uronic acid residues, i.e. 2-deoxy-2-sulphamido-D-glucopyranose 6-sulphate and L-idopyranosyluronic acid 2-sulphate. 13C NMR has been used to study the interactions of heparinase-derived and purified heparin disaccharide with N- and C-terminally-blocked tripeptides GRG and GKG. Titration of the disaccharide with peptide indicates that GRG binds the disaccharide more strongly than does GKG, with interactions in either case being stronger at uronate ring positions. In the presence of GRG, a carboxylate pKa depression suggests electrostatic interactions between the arginine guanidinium group and the uronate carboxylate group. 13C relaxation data have been acquired for all disaccharide and peptide carbons in the presence and absence of GRG and GKG. 13C relaxation rates for the disaccharide are significantly faster in the presence of peptide, especially with GRG. Analysis of these relaxation data has been done in terms of molecular diffusion constants, D and D, and an angle α between D and a molecular frame defined by the moment of inertia tensor calculated for an internally rigid disaccharide. Disaccharide conformational space in these calculations has been sampled for both uronate half-chair forms (2H1 and 1H2) and over a range of glycosidic bond angles defined by motional order parameters and inter-residue nuclear Overhauser effects (±30° from the average). In the absence of peptide, the ratio D/D falls between 0.4 and 0.7; therefore molecular diffusion occurs preferentially about D, which runs through both disaccharide rings. In the presence of peptide, D/D is decreased, indicating that GRG is oriented along D and proximal to the uronic acid ring. A model for this is shown.

Original languageEnglish (US)
Pages (from-to)447-454
Number of pages8
JournalBiochemical Journal
Volume315
Issue number2
DOIs
StatePublished - Apr 15 1996

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