Takeshi Hashizume

Conformational Rigidity of N4-Acetyl-2′-O-methylcytidine Found in tRNA of Extremely Thermophilic Archaebacteria (Archaea)

Abstract The conformational characteristics of N 4-acetyl-2′-O-methylcytidine (ac4Cm), which is one of the modified cytidines unique to the tRNA of extremely thermophilic archaebacteria, and related nucleosides, N 4-acetylcytidine (ac4C), 2′-O-methylcytidine (Cm) and cytidine, were analyzed by proton nuclear magnetic resonance spectroscopy. Ribose methylation and N 4-acylation were found to confer high conformational rigidity to the ribose moiety, suggesting that these post-transcriptional modifications play a role in structural stabilization of tRNA, which is of particular importance at high temperature.

Conformatzonal Properties of a Novel Modified Nucleoside, 5-Formylcytidine, Found at the First Position of the Anticodon of Bovine Mitochondrial tRNAMet

Abstract Conformational properties of a novel modified nucleoside, 5-formylcytidine (f5C), which is found at the first position of the anticodon of bovine mitochondrial tRNAMet, were analyzed by 1H-NMR spectroscopy. f5C has a normal amino tautomeric form at position 4 of the base moiety. The results indicate the presence of an intramolecular hydrogen bond between the carbony1 of the 5-formyl group and the 4-amino function. f5C was found to exhibit the C3′-endo conformation exclusively and the enthalpy difference (ΔH) between the C2′-endo and C3′- endo forms was found to be 1.56 ± 0.13 kcal/mol, indicating f5C to be one of the most conformationally rigid nucleosides yet analyzed. The conformational rigidity of f5C may contribute to regulation of codon recognition by tRNAMet.

Structural characterization of four ribose-methylated nucleosides from the transfer RNA of extremely thermophilic archaebacteria

Structures of the ribose-methylated nucleosides 5,2′-O-dimethylcytidine (1), N4-acetyl-2′-O-methylcytidine (2), 2-thio-2′-O-methyluridine (3), and N2,N2,2′-O-trimethylguanosine (4) from the transfer RNA of Sulfolobus solfataricus, Thermoproteus neutrophilus, and Pyrodictium occultum have been established, and verified by chemical synthesis.

Applications of LC/MS and Tandem Mass Spectrometry to the Characterization of Nucleosides in Mixtures

Abstract Liquid chromatography-mass spectrometry (LC/MS) and tandem mass spectrometry (MS/MS) provide new approaches for structural studies of nucleosides, in the nanogram range, in mixtures. Examples are given of the use of LC/MS for rapid screening of synthesis reaction mixtures, and of MS/MS for the detection and characterization of nucleoside isomers in RNA hydrolysates.