J. D. Cutnell

Systematic elimination of RF pulse defect errors in Fourier transform spin-lattice relaxation measurements

Abstract The elimination of rf pulse defect errors in Fourier transform spin-lattice relaxation measurements is discussed. The standard 180°−τ−90°−t pulse sequence is used. However, the phase of the 90° pulse is shifted by 180° on every repetition of the sequence, while the phase of the 180° pulse is unaltered. Implementation of this phase shifted pulse sequence is discussed for a JEOL PFT-100 single phase detector spectrometer.

Proton magnetic resonance characterization of the dynamic stability of the heme pocket in myoglobin by the exchange behavior of the labile proton of the proximal histidyl imidazole.

The assigned exchangeable proton signals in the proton nuclear magnetic resonance spectra of sperm whale deoxy and Met-cyano myoglobin in H2O solution were found to exhibit pH-dependent saturation transfer from the bulk water, which allowed determination of the kinetics and mechanism of the labile proton exchange with solvent. The exchange rates are base catalyzed for both protein forms, with the rate eight times faster in Met-cyano than in deoxy myoglobin. The exchange rate is taken as a measure of the magnitude of the fluctuation in the protein conformation near the heme cavity. On the basis of tritium exchange methods, the greater stability of the unligated relative to the ligated state in myoglobin has also been reported for hemoglobin. The present study, however, localizes the differential kinetic stability on the F helix whose flexibility has been implicated in the mechanism of cooperativity. The observation that filling the hydrophobic vacancy on the proximal side of the heme near the proximal histidine in Met-cyano myoglobin wih cyclopropane increases the proton lability argues against the role for this hole in facilitating the flexibility of the F helix in the native protein.

NONDIPOLAR CONTRIBUTIONS TO 13C RELAXATION IN MOLECULES

Our laboratory has been concerned with the determination of the microdynamics of peptide spatial motions in solution.’q2 The information we seek is concerned with whether or not peptide hormones in solution, and interacting with binding macromolecules, are rigid units. The alternative is that in either or both cases the molecules possess internal degrees of freedom, the knowledge of which might give us some insight into the forces that hold these entities together topologically and that are thought to be responsible for their specificity and actions. In addition, knowledge of the flexibility of the molecules should tie in with the considerable amount of effort going into theoretical studies of peptide structure. For example, the microdynamic activation energies for segmental internal rotation should be related to the bamers between local potential wells in the phi-psi theoretical conformational plots. Nuclear magnetic resonance (nmr) relaxation techniques are the only methods now available for accurately determining the microdynamic parameters of molecules such as these peptides without introducing perturbing “labels.” For this reason many nmr laboratories in the world are engaged in such studies. The nuclei commonly studied are: protons, deuterons, and carbon-1 3. The observable is, in each case, called the spin-lattice relaxation time (TI ) and what is desired is a relation between this experimental parameter and the microdynamic one called the molecular reorientation time (T,). This latter can be conveniently thought of as the characteristic time during which a molecule’s rotational coordinates remain constant between jumps to new positions due to random collisions with solvent molecule^.^ For reasons connected with theoretical difficulties in forming a satisfactory relation between experimental proton Tl’s and T,, proton measurements are not used for these purposes at the present time. Of the remaining nuclei, deuteron relaxation provides the most direct way of determining T,, but involves lengthy chemical syntheses of specifically labeled compounds. On the other hand, natural abundance carbon-1 3 spectroscopy requires no synthetic work but unfortunately for this case the theoretical interpretation of the results is less clear. This may be pointed out as follows. Equation 1 gives the relation between