HCCH-type experiments

In fully C-labeled proteins, the side chains of amino acids can be assigned using CCH or HCCH-TOCSY-type experiments (Fesik and Zuiderweg, 1990 Fesik et al., 1990 Bax et al., 1990b Kay et al., 1993). The three-dimensional HCACO-TOCSY experiment (Kay et al., 1992) is an extension of these experiments that uses the carbonyl chemical shifts to separate overlapping resonances. 

Exclusive TACSY (E.TACSY) mixing sequences (see Section X.E) for aliphatic-selective C- C Hartmann-Hahn transfer without perturbing the polarization of carbonyl C spins (Schmidt et al., 1993 Weisemann et al., 1994 Abramovich et al., 1995) have been used in HCCH-E.COSY-type experiments (see Fig. 39) for the measurement of V(C, H ) coupling constants (see Fig. 40). 

To overcome the problem of low dispersion of sugar resonances in nucleotides Hu et al. proposed to incorporate HCN transfer into HCCH-type sequences resulting in 3D MQ-HCN-CCH-TOCSY and 3D MQ-HCN-CCH-COSY experiments. The coherence transfer pathway starts with HT-CT transfer followed by an out-and-back transfer to N1/N9 for indirect detection and subsequent transfer through network. As a result the CCH spectra are additionally resolved with chemical shift. During the C -N1/N9 transfer MQ HT/CT coherence is active to reduce the relaxation losses due to relaxation. The experiments were successfully applied to a 23-mer RNA aptamer. 

Clearly the homonuclear and the heteronuclear experiments could be combined in the reverse order i.e. HSQC-NOESY and HSQC-TOCSY. The main advantage in these schemes relates to N-edited experiments in which the narrower amide proton spectral width is sampled during f and the full proton spectral width is collected during t. On the other hand water suppression is more effective when HSQC follows NOESY. Also the sensitivity enhancement can be incorporated into the NOESY-HSQC experiment. Eor the TOCSY-HSQC or HSQC-TOCSY it does not matter because both the TOCSY and HSQC sequences can be implemented with the sensitivity enhancement. It should be mentioned that the TOCSY type of transfer is more effective between C nuclei than between protons and therefore the HCCH-TOCSY experiment is preferred when a doubly labeled sample is available. 

Another report on NMR studies of perdeuterated proteins by Agarwal et al. has shown that the small amount of residual protonation is sufficient to perform 2D and 3D MAS solid-state NMR experiments. In particular, a HCCH-TOBSY type experiment was suggested, which was successfully employed to assign the methyl resonances in aliphatic side chains in a perdeuterated sample of the SH3 domain of chicken a-spectrin. 

Photolysis of H3NBH3 with 121.5 nm radiation yields imidoborane, HBNH, which has been of theoretical interest Spectral shifts observed for several isotopic species containing °B, N, and D show clearly that the spectrum is due to HNBH which is isoelectronic with HBO, HCN and HCCH. From the spectrum of the isolated species two of the and one of the tr-type vibration frequencies for a linear molecule have been obtained. The location of the missing S (B-H stretch) frequency has been calculated. A comparison of observed and calculated frequencies for HBNH is given in Table 7. Another isolated product observed in these experiments is identified as HNB. This radical may be generated by photodissociation of HNBH subsequent to its formation. In this respect the photolysis mechanism would be similar to the formation of C2H from acetylene. 

The transfer of an H atom from one site to another, as in the HCN — NCH isomerization, can be viewed as a special type of internal rotation. A hindered internal rotor treatment of such motions was found [148, 149] to yield an increase in the reactant state density by a factor of 3 to 4 for both HCN and HCCH at the thresholds for CH bond dissociations. Furthermore, for HCN, where the dissociation energy is well known, the resulting low pressure limit rate coefficients were found to be in much improved agreement with experiment. This study also provided a simple general formula for estimating the effect of such corrections for arbitrary isomerizations (Eq. (2.31) in [149]). Illustrative calculations suggested that such effects may be important even in larger molecules. 

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