Backbone dynamics of the major tacrolimus (FK506) binding protein (FKBP-12, 107 amino acids) have been studied using 15N relaxation data derived from proton-detected two-dimensional 1H-15N NMR spectroscopy. 15N spin-lattice relaxation rate constants (R1), spin-spin relaxation rate constants (R2), and heteronuclear NOEs were determined for over 85% of the backbone amide 15N nuclei. A model free formalism [Lipari, G., & Szabo, A. (1982) J. Am. Chem. Soc. 104, 4546-4559; Lipari, G., & Szabo, A. (1982) J. Am. Chem. Soc. 104, 4559-4570] was used to derive values for the generalized order parameter (S2), the effective correlation time for internal motions (tau e), and the chemical exchange line width (Rex) for each N-H bond vector. The final optimized overall correlation time (tau m) was 9.2 ns. The average order parameter (S2) describing the amplitude of motions on the picosecond time scale was found to be 0.88 +/- 0.06. Motions on the picosecond time scale are restricted at the N and C termini, consistent with previous NMR structural studies indicating well-defined beta-strands in these regions. With the exception of the flap region from resides 82 to 87, no regions appear to be significantly disordered on the picosecond time scale. Residues in several regions of the protein exhibit high Rex terms, indicating possible motions on the millisecond to microsecond time scale due to chemical exchange and/or conformational averaging effects. Possible effects of tacrolimus (FK506) binding on FKBP-12 dynamics are discussed in the context of previously determined solution structures for FKBP-12 in the uncomplexed [Michnick et al. (1991) Science 252, 836-839; Moore et al. (1991) Nature 351, 248-250] and complexed [Meadows et al. (1993) Biochemistry 32, 754-765] states.

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Cardiotoxin analogue II (CTX II) is an all beta-sheet, small molecular mass (6.8 kDa), basic protein possessing a wide array of biological properties. Nearly complete assignment of the protonated carbon resonances has been achieved by heteronuclear NMR experiments. The study shows that the correlation between the carbon-13 chemical shifts and CTX II structure is good in general, but interesting deviations are also noticed. To characterize the internal dynamics of CTX II, longitudinal, transverse relaxation rates and heteronuclear 13C{1H} NOEs were measured for alpha-carbons at natural abundance by two-dimensional NMR spectroscopy. Relaxation measurements were obtained in a 14.1 T spectrometer for 50 residues, which are evenly spread along the CTX II polypeptide chain. Except for five alpha-carbons, all data were analyzed from a simple two-parameter spectral density function using the model free approach of Lipari and Szabo. The microdynamical parameters (S2, taue, and Rex) were calculated with an overall rotational correlation time (taum) for the protein of 4.8 ns. For most residues, the alpha-carbons exhibit fast (taue < 30 ps) restricted libration motions (S2 = 0.79-0.89). The present study reveals that the functionally important residues located at the tips of the three loops are flexible, and the flexibility of residues in this region could be important in the binding of cardiotoxins to their putative "receptors" which are postulated to be located on the erythrocyte membrane. In addition, the results obtained in the present study support the earlier predictions on the relative role of the lysine residues in the erythrocyte lytic activity of cardiotoxins.