5S rRNA modification in the hyperthermophilic archaea Sulfolobus solfataricus and Pyrodictium occultum.

Abstract

The 5S rRNAs from Sulfolobus solfataricus and Pyrodictium occultum were digested to nucleosides and analyzed using directly-combined HPLC/mass spectrometry. P. occultum 5S rRNA contains two modified nucleoside species, N4-acetylcytidine (ac4C) and N4-acetyl-2'-O-methylcytidine (ac4Cm). Oligonucleotides were generated from P. occultum 5S rRNA by RNase T1 hydrolysis, and their molecular weights were determined using electrospray mass spectrometry and compared with those predicted from the P. occultum 5S RNA gene sequence. Deviation in mass between expected and observed molecular weights permitted ac4Cm to be located at position 35, in the nonanucleotide CAA-CACC[ac4Cm]G, and the ac4C in one or both of two (C,U)G trinucleotides. 2'-O-Methylcytidine is unambiguously characterized in S. solfataricus 5S rRNA, confirming earlier tentative assignments at the analogous sequence position (Stahl, D.A., Luehrsen, K.R., Woese, C.R., and Pace, N.R. (1981) Nucleic Acids Res., Vol. 9, pp. 6129-6137; Dams, E., Londei, P., Cammarano, P., Vandenberghe, A., and De Wachter, R. (1983) Nucleic Acids Res. Vol. 11, pp. 4667-4676). Potential effects of the presence of ac4C and ac4Cm on thermal stabilization of 5S rRNA in thermophiles are discussed.

A Gateway platform for functional genomics in Haloferax volcanii: deletion of three tRNA modification genes

“…This modification is found at position 34 in five H. volcanii tRNAs (Gupta 1984, Gupta 1986), but its function is unknown. It has been postulated that the conformational rigidity of this nucleoside could stabilize RNA molecules of thermophilic archaea (Kawai et al 1992, Bruenger et al 1993, Oliva et al 2007). Our data indicate that an H. volcanii strain with tRNA lacking ac 4 C remains viable.…”

Conformational Studies of 4‐N‐Carbamoyldeoxycytidine Derivatives and Synthesis and Hybridization Properties of Oligodeoxyribonucleotides Incorporating these Modified Bases

“…13 C NMR spectra were recorded at 68 MHz and the chemical shifts were measured from the solvent peak as an internal standard.…”

“…Supporting Information (see footnote on the first page of this article): Tautomerism studies of 4-N-carbamoyldeoxycytidine derivatives by use of 13 C NMR chemical shifts.…”

“…[1,2] The 1 H NMR analysis and the ab initio MO calculations revealed that these 4-N substituents were fixed in the same "proximal" geometry as that reported in the case of 4-N-acetylcytidine found in tRNA [5][6][7][8][9][10][11] and rRNA. [12,13] From the crystal [14] and NMR [15] structures reported to plementary strands without disturbing the structure of DNA duplexes, hence changing the orientation of the carbamoyl group in such a manner that a stable Watson-Crick base pair can be formed with the guanine base at the opposite site. It should be noted that the base recognition ability (G against T, C and A) of these modified cytosine bases can be preserved satisfactorily.…”

The single pseudouridine residue inEscherichia coli16S RNA is located at position 516

“…These are nucleosides methylated either on the base or on the 2'-hydroxyl of ribose and pseudouridine (I, 5-(3-D-ribofuranosyluracil), the 5-ribosyl isomer of uridine, and its derivatives, such as l-methyl-3-(3-amino-3-carboxypropyl) pseudouridine in eukaryotic small subunit rRNA (5,6). Other modified nucleosides in rRNA not included in these categories are dihydrouridine in E.coli 23S rRNA (7,8) and N4-acetylcytidine (9, 10) and its 2'-O-methyl derivative (10) in other rRNAs.…”