Molecular dynamics and structure of the random coil and helical states of the collagen peptide, alpha 1-CB2, as determined by 13C magnetic resonance.
Carbon-13 chemical shifts, spin-lattice (T1) and spin-spin (T2) relaxation times, and 13C-[1H] nuclear Overhauser enhancements (NOE) have been determined for the random coil and triple helical states of the alpha 1-CB2 fragment of rat skin collagen. Assignment of all aliphatic resonances of this 36 residue peptide in the random coil state (30 degrees) has been achieved with the aid of model polypeptides containing pyrrolidine residues. The chemical shifts and intensities of the Pro and Hyp C-gamma resonances show that (see article) 90% of the X-Pro and X-Hyp bonds are trans in both helix and coil conformations. From T1 measurements rotational correlation times (tau-eff) of ca. 0.45 nsec are calculated for interior C-alpha carbons in the coil, while taueff values of the side chain and near terminal carbons are found to be 2-9 times smaller. These results along with the narrow natural line widths (3-5 Hz) and maximal NOE values (2.8 plus or minus 0.3) demonstrate the high degree of backbone mobility, due to segmental motion, in the unordered state of the peptide. By contrast, the broad lines (50-90 Hz) and small NOE values (1.3 plus or minus 0.3) for the alpha carbons in the helical state (2 degrees) suggest much slower motion. The line widths and NOE values together with the C-alpha T1 values (0.025-0.040 sec) correspond to correlation times which are in reasonable agreement with those calculated for an axially symmetric rigid ellipsoid, undergoing rotational diffusion, having dimensions approximating those of a collagen-type triple helical aggregate of three alpha 1-CB2 chains. A satisfactory computer simulation of the experimental 2 degrees spectrum is obtained by assigning the narrow aliphatic resonances in the spectrum (line widths 5-40 Hz) to (a) carbons in the small amounts of alpha 1-CB2 (3 mol %) and alpha 1-CB1 (2.5 mol %) random coil conformations, (b) carbons in the flexible terminal triplets of the helix, and (c) Ala, Leu, and Phe methyl and phenyl carbons. The side chain carbon line widths obtained from the simulation--when compared with side chain line widths calculated for a rotating rigid ellipsoid with internal motion--indicate rapid axial reorientation of methyl and phenyl groups. With the exception of the Hyp residue the line widths suggest local motion for at least some carbons in most other side chain moieties. The Hyp C-beta and C-gamma line widths indicate the presence of little if any rapid Hyp ring motion.
D000818:Animals; D002247:Carbon Isotopes; D002852:Chromatography, Ion Exchange; D003094:Collagen; D003201:Computers; D005583:Fourier Analysis; D009682:Magnetic Resonance Spectroscopy; D008433:Mathematics; D010446:Peptide Fragments / Q000032:analysis; D011487:Protein Conformation; D051381:Rats; D012867:Skin; D013997:Time Factors
D002247:Carbon Isotopes; D010446:Peptide Fragments; D003094:Collagen