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

Abstract

We have determined the amplitude of nanosecond fluctuations of the collagen azimuthal orientation in intact tissues and reconstituted fibers from an analysis of 13C NMR relaxation data. We have labeled intact rat calvaria and tibia collagen (mineralized and cross-linked), intact rat tail tendon and demineralized bone collagen (cross-linked), and reconstituted lathyritic (non-cross-linked) chick calvaria collagen with [2-13C]glycine. This label was chosen because one-third 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 degrees C. The measured NMR parameters have been 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. We have shown that if root mean square fluctuations in the azimuthal orientations are small, gamma rms much less than 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. Our analysis shows that, at 22 degrees C, tau is in the 1-5-ns range for all samples and gamma rms is 10 degrees, 9 degrees, and 5.5 degrees for the non-cross-linked, cross-linked, and mineralized samples, respectively. At -35 degrees C, gamma rms is less than 3 degrees for all samples. These results show that mineral and low temperature significantly restrict the amplitude of nanosecond motions of the collagen backbone.(ABSTRACT TRUNCATED AT 250 WORDS)

Water scaffolding in collagen: Implications on protein dynamics as revealed by solid‐state NMR

“…In line with previous reports, [11][12][13][14][17][18][19][20]59-62 our solid-state NMR studies of collagen samples of various origin confirm that the amplitude of collagen backbone and sidechain motions increases significantly on increasing the water content. This conclusion is supported by the changes observed in three different in nature NMR observables: (i) the linewidth dependence on the 1 H decoupling frequency; (ii) 13 C CSA changes for the peptide carbonyl groups, and (iii) 1 H- 13 C dipolar dephasing rates.…”

“…Despite the simplicity of the model considered, these results suggest that the increase of the water content could lead to significant increase of the internal restricted rotations (i.e., librations) about C-C a or N-C a bonds of the collagen backbone. Overall, the observed dependence of collagen dynamics on the water content is in agreement with the previous reports, [11][12][13][17][18][19][20][59][60][61][62] as well as the results of studies on the effect of hydration of the molecular mobility of collagen in bone and cartilage. [18][19][20][59][60][61][62] The dependence of collagen dynamics on the water surrounding appears to be its intrinsic property, which is retained in different environments in the presence of lipids, calcium phosphate, glycosaminoglycans, and other species.…”

“…It has been shown that the triple-helical structure causes more ordering of the hydration water than the denaturated single polipeptide chains. 10 The molecular dynamics of the collagen backbone in intact connective tissues has been investigated using 13 C NMR line shape analysis to measure 13 CO chemical shift anisotropy (CSA) by Torchia et al 11 They used [1][2][3][4][5][6][7][8][9][10][11][12][13] C]glycine to label samples from various sources in order to investigate the effect of crosslinking and mineralization on the peptide backbone motion in the collagen fibril. In another report by Torchia et al, 12 the amplitude of motional fluctuations in collagen in intact tissues and reconstituted fibres were determined using 13 C NMR relaxation time measurements.…”

“…In both cases, 11,12 it was assumed that protein backbone motions are primarily a consequence of reorientation about the long axis of the molecule. This motional model of a collagen rod librating about its helix axis was questioned in reference [13]; since, considering the size of the collagen molecule and the presence of crosslinked molecules, motional amplitudes derived for the helix axis libration (up to 33 ) were unusually high.…”

A high‐resolution solid‐state 13C‐NMR study on crystalline bovine heart cytochrome‐c oxidase and lysozyme

“…Similarly, a short T1 of Gly Ca was previously noted for calf skin collagen (4.8 s; Sait6 and Yokoi, 1992). This was previously explained in terms of the presence of collagen azimuthal fluctuation as 15 -10 O, with a correlation time of 1 -5 ns (Sarkar et al, 1985). Therefore, it should be pointed out that the backbone of cytochrome-c oxidase fluctuates in the crystal much more than in lysozyme.…”