ISSN:
1573-5001
Keywords:
Collagen
;
Kunitz domain
;
Multiple conformations
;
Exchange rate
Source:
Springer Online Journal Archives 1860-2000
Topics:
Biology
,
Chemistry and Pharmacology
Notes:
Summary The human α3-chain type VI collagen C-terminal Kunitz domain fragment (α3(VI)) has been studied by two-dimensional 1H−1H and 1H−13C NMR spectroscopy at 303 K. It is shown that the secondary structure of the protein is strikingly similar to that of BPTI, and that a number of unusual Hα chemical shifts, which are highly conserved in Kunitz-domain proteins, are also observed for α3(VI). Further-more a series of exchange cross peaks observed in 1H−1H spectra shows that a large number of protons in the central β-sheet exist in two different chemical environments, corresponding to two unequally populated conformations that are slowly exchanging on the NMR time scale. Several protons, including Ser47(53) Hα, Arg32(38) Hγ2, and Gln48(54) Hβ2, all located in the vicinity of the Trp21(27) ring in the crystal structure of α3(VI) [Arnoux, B. et al. (1995) J. Mol. Biol., 246, 609–617], have very different chemical shifts in the two conformations, the most affected being Gln48(54) Hβ2 (Δδ=1.53 ppm), which is placed directly above the Trp21(27) ring in the crystal structure of α3(VI). It is concluded that the origin of the multiple conformations of the central β-sheet is a reorientation of the Trp21(27) ring. From the intensities of corresponding signals in the two conformations, the population of the minor conformation was found to be 6.4±0.2% of that of the major conformation, while a rate constant kM=1.01±0.05 s-1 for the major to minor interconversion was obtained from a series of NOESY spectra with different mixing times. In addition, it is shown that Cys14(20)-Cys38(44) disulfide bond isomerization, previously observed in BPTI [Otting, G. et al. (1993) Biochemistry, 32, 3570–3582], is also likely to occur in α3(VI).
Type of Medium:
Electronic Resource
URL:
http://dx.doi.org/10.1007/BF00228142
Permalink
