13C LABELED GLYCEROL

University Of Calif-Los Alamos Nat Lab, Los Alamos NM

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Abstract

The SIR provided [13C, 1,3-'H4 ]Glycerol; 14g Uniform 13C enrichment has become widely used for resonance assignment studies of proteins and, more recently, nucleic acids. Along with the various benefits, there are several limitations to this labeling pattern which stem from the presence of one bond 13C-13C couplings. In addition to the degraded reslution in the 13C dimension which can only partially be alleviated by constant time techniques, no inderct detected 13C relaxation experiments have been developed which cercumvent the artifacts due to the 13C-13C scalar interactions. The alternating carbon enrichment procedure we have developed for E. coli expression systems allows for both relaxation experiments as well as various 13C editing assignment experiments. This method involves culturing the organisms using [2-13C]- or [1,3-13C21glycerol as a carbon source. Furthermore, we have demonstrated that by combining random fractional deuteration with selective 13C enrichment, one can analyze lH detected relaxation of methylene carbons which has heretofore not been feasible. As a result by using the two opposite patterns of 13C enrichment it should be possible to deterine the motional parameters of nearly all sites of a protein or nucleic acid molecule. In addition, particularly in the case of nucleic acids, we anticipate that this approach to selective labeling will offer a substantial complement to structural studies via uniform 13C enrichment.