Dynamics of collagen in articular cartilage studied by solid-state NMR methods.

Abstract

Methods for studying the fast molecular dynamics of the rigid macromolecules in cartilage are described. The strong dipolar couplings and chemical shift anisotropies of these molecules necessitate application of solid-state nuclear magnetic resonance (NMR) techniques such as magic-angle spinning, cross-polarization, and high-power dipolar decoupling to obtain resolved NMR spectra. The molecules in cartilage that are amenable to these techniques are collagen and the rigid portion of the glycosaminoglycans (mostly hyaluronan). Site-specific mobility information is obtained from scaled dipolar couplings measured in 2D NMR experiments. Motionally averaged dipolar couplings can be interpreted in terms of order parameters that provide information about the amplitudes of molecular motions. Qualitative dynamics information is obtained from the simple wideline separation experiment measuring 1H-1H widelines representing the strength of the 1H-1H dipolar coupling. Quantitative values for molecular order parameters are obtained from precise measurements of 1H-13C dipolar couplings along the C-H bond vector. Two experimental techniques, the Lee-Goldburg cross-polarization and the dipolar coupling/chemical shift experiment, are illustrated to measure these 1H-13C dipolar couplings. Unlike glycosaminoglycans in cartilage, the collagen moiety remains substantially ordered, undergoing fast small amplitude motions. As enzymes cleave the macromolecules in articular cartilage in the course of arthritis, solid-state NMR techniques are capable of characterizing the increased motions of their degradation products in diseased cartilage.