Production of specific site probes of tRNA structure by enrichment with carbon 13 at particular locations
Open Access
- 10 October 1979
- journal article
- research article
- Published by Oxford University Press (OUP) in Nucleic Acids Research
- Vol. 7 (3) , 765-779
- https://doi.org/10.1093/nar/7.3.765
Abstract
Escherichia coli C6 rel met cys was cultured in a stringently defined minimal medium containing 13C-enriched metabolites in order to (1) achieve maximal 13C isotopic enrichment of tRNA; and (2) produce site specific but natural, non-perturbing NMR probes of tRNA structure and function. Growth conditions were manipulated to achieve optimal culture growth concomitant with maximal in vivo incorporation of various 13C-enriched nucleic acid precursors, including L-[methyl-13C] methionine, [2-13C] adenine, and [2-13C] uracil. Effective blockage of purine biosynthesis de novo was accomplished with the addition of the antimetabolite 6-mercaptopurine to the growth medium. Transfer RNAs specifically 13C-enriched in all methyl groups (57 atom %), C2 of adenine (60 atom %), and C2 of uracil (82 atom %) and C2 of cytosine (73 atom %) have been produced.Keywords
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