Proteins deuteration

Blood Fluoride reactivation method Phospho-rylated BuChE (+ other proteins) GC-MS GC-HR-MS With large volume injection Phosphonofluo- ridates i.s. deuterated OP or plasma exposed to deuterated OP Easily accessible internal standards and reference compounds LOD 10 pg/ml (0.05-0.1% BuChE inhibition) Not applicable to all OP s... 

Although the first NMR spectra of deuterated proteins were published already in 1968, the deuteration was mainly employed for the purpose of simplifying one-dimensional proton spectra.53,54 Later in the 1980s, LeMaster and co-workers employed random fractional deuteration for the assignment of thioredoxin using homonuclear two-dimensional spectroscopy.55 However, it was realized that deuteration significantly improves the spectral quality of homonuclear multidimensional NMR experiments. 

The effect of deuteration, for the needs of sequence-specific resonance assignments using triple-resonance experiments, was first demonstrated by Bax and co-workers on the 19.7 kDa protein calcineurin B in 1993.56 By replacing the H spin with deuterium, the transverse relaxation time of 13C spin is increased by nearly an order of magnitude due to the 6.5 times smaller gyromagnetic ratio of 2H in comparison to in.52,56 Not surprisingly, deuteration was utilized for the aid of structure determination of several... 

Size restrictions are again similar to that encountered in protein NMR up to ca. 20-30 kDa complex size, line widths will usually be acceptable to allow detection of the XH resonances of the labeled or unlabeled component, including NOESY transfer steps. Beyond that, deuteration of the protein component becomes essential to reduce excessive line broadening. For the observation of the (15N-) labeled component (i.e., isotope editing), the use of TROSY techniques will further extend the size limit (Chapt. 10) however, this approach does not work for unlabeled components. 

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