Nanosecond fluctuations of the molecular backbone of collagen in hard and soft tissues: a carbon-13 nuclear magnetic resonance relaxation study

Abstract

The amplitude of nanosecond fluctuations of the collagen azimuthal orientation in intact tissues and reconstituted fibers from an analysis of 13C NMR relaxation data. Intact rat calvaria and tibia collagen (mineralized and cross-linked), intact rat tail tendon and demineralized bone collagen (cross-linked), were labeled and lathyritic (non-cross-linked) chick calvaria collagen was reconstituted with [2-13C]glycine. This label was chosen because 1/3 of the amino acid residues in collagen are glycine and because the 1H-13C dipolar coupling is the dominant relaxation mechanism. Spin-lattice relaxation times (T1) and nuclear Overhauser enhancements were measured at 15.09 and 62.98 MHz at 22 and -35.degree. C. The measured NMR parameters were analyzed by using a dynamic model in which the azimuthal orientation of the molecule fluctuates as a consequence of reorientation about the axis of the triple helix. If root mean square fluctuations in the azimuthal orientations are small, .gamma.rms .mchlt. 1 rad, the correlation function decays with a single correlation time .tau. and T1 depends only upon .tau. and .gamma.rms and not the detailed model of motion. At 22.degree. C, .tau. is in the 1-5-ns range for all samples and .gamma.rms is 10.degree., 9.degree. and 5.5.degree. for the non-cross-linked, cross-linked and mineralized samples, respectively. At -35.degree. C, .gamma.rms is less than 3.degree. for all samples. Mineral and low temperature significantly restrict the amplitude of nanosecond motions of the collagen backbone. Similar conclusions were obtained from an analysis of [1-13C]glycine line shapes, except that the .gamma.rms values reported were 3-4 times larger than .gamma.rms values reported herein. This difference in .gamma.rms values apparently is obtained because line shapes are sensitive to much slower motions than are relaxation parameters.