Partial assignment of resonances in the 19F nuclear magnetic resonance spectra of 5-fluorouracil-substituted transfer RNAs.
Features of the 19F nuclear magnetic resonance (NMR) spectra of three purified 5-fluorouracil-(FUra-) substituted Escherichia coli tRNAs, tRNA(1Val), tRNA(mMet), and tRNA(fMet), are compared. Each of the tRNA species can be resolved into two isoaccepting forms, A and B, whose 19F NMR spectra differ in the shift of one peak from the 4.5 to 4.8 parts per million (ppm) range (FUra = O) in the spectrum of isoacceptor B upfield to ca. -15 ppm in that of isoacceptor A. Because the sequences of the two isoacceptors of each tRNA differ only at one position in the D loop, that normally occupied by a dihydrouridine residue, we assign the 4.5 ppm peak in the spectrum of fluorine-labeled tRNA(1Val) to FUra17 and the resonance at 4.6 ppm in the spectrum of fluorouracil-substituted tRNA(mMet) to FUra20. A reciprocal 19F[19F] nuclear Overhauser effect is observed between the downfield peaks A and B in the 19F NMR spectrum of 19F-labeled tRNA(1Val). Assuming that fluorine-labeled tRNA(1Val) has a structure similar to that of yeast tRNA(Phe), only FUra54 and -55 are close enough (4-5 A) to give an appreciable 19F homonuclear Overhauser effect. Peaks A and B have therefore been assigned to FUra54 and -55. As the temperature is raised from 30 to 45 degrees C, the intensity of peak B (6.6 ppm) in the spectrum of 19F-labeled tRNA(1Val) gradually shifts upfield to 6.4 ppm (Tm = 36 degrees C), indicating a temperature-dependent slow exchange of the corresponding 5-fluorouracil residue between two magnetically distinct environments.(ABSTRACT TRUNCATED AT 250 WORDS)
Hardin
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CC
Department of Biochemistry and Biophysics, Iowa State University, Ames 50011.
Gollnick
P
P
Horowitz
J
J
D004926:Escherichia coli / Q000235:genetics; D005472:Fluorouracil; D008274:Magnesium / Q000494:pharmacology; D009682:Magnetic Resonance Spectroscopy / Q000379:methods; D009690:Nucleic Acid Conformation / Q000187:drug effects; D009994:Osmolar Concentration; D012345:RNA, Transfer, Amino Acid-Specific / Q000302:isolation & purification; D012346:RNA, Transfer, Amino Acyl / Q000302:isolation & purification; D012358:RNA, Transfer, Met; D012366:RNA, Transfer, Val / Q000302:isolation & purification; D012965:Sodium Chloride / Q000494:pharmacology
D012345:RNA, Transfer, Amino Acid-Specific; D012346:RNA, Transfer, Amino Acyl; D012358:RNA, Transfer, Met; D012366:RNA, Transfer, Val; C026464:tRNA(m)(Met), methionine-; C024988:tRNA, formylmethionine-; D012965:Sodium Chloride; D008274:Magnesium; D005472:Fluorouracil