Procedure for C2 deuteration of nucleic acids and determination of A psi 31 pseudouridine conformation by nuclear Overhauser effect in yeast tRNAPhe.
Nuclear Overhauser effect (NOE) combined with semispecific deuteration provides a general strategy for identification of exchangeable protons in nucleic base pairs, and has been extended to NOEs involving purine C2 protons in tRNA. Deuterated tri-ethyl orthoformate was condensed with 5(4)-amino imidazole 4(5)-carboxamide to yield C2 deuterated hypoxanthine. C2 deuterated hypoxanthine was fed to a purine requiring mutant of yeast and C2 deuterated yeast tRNAPhe was isolated. This C2 deuterated tRNAPhe was used to identify A psi 31 and U8-A14. A psi 31 was found to be bonded through N1H. The utility of C2 deuteration in nucleic acid NMR is thus demonstrated.
D001483:Base Sequence; D003903:Deuterium; D009682:Magnetic Resonance Spectroscopy / Q000379:methods; D009154:Mutation; D009690:Nucleic Acid Conformation; D011560:Pseudouridine / Q000032:analysis*; D012346:RNA, Transfer, Amino Acyl / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000235:genetics; D014529:Uridine / Q000031:analogs & derivatives*
D012346:RNA, Transfer, Amino Acyl; D011560:Pseudouridine; D003903:Deuterium; D014529:Uridine