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P F Crain

Publications and source records attributed to P F Crain.

At least 55 records · Page 3Linked to original sources
PubMed1987

Isolation and structure elucidation of an epoxide derivative of the hypermodified nucleoside queuosine from Escherichia coli transfer RNA.

A new nucleoside has been identified in tRNATyr from Escherichia coli MRE 600, where it replaces the highly modified nucleoside queuosine. The nucleoside is also present in a large amount relative to queuosine in mixed tRNA from E. coli strains MRE 600 and W (from which it was isolated for characterization). The new nucleoside has been characterized as an epoxy derivative of queuosine: 7-(5-[(2,3-epoxy-4,5-dihydroxycyclopent-1-yl)amino]methyl)-7-de azaguanosine, oQ, based on data from directly combined liquid chromatography/mass spectrometry, high resolution mass spectrometry, and proton NMR spectroscopy. Nucleoside oQ is also present in small amounts in mixed tRNA from E. coli B. Isomerization of oQ occurs readily under alkaline conditions to give a rearranged product, oQ', characterized as 7-(5-[(3,4-epoxy-2,5-dihydroxycyclopent-1-yl)amino]methyl)-7-deaza guanosine. The present finding constitutes the first report of epoxide formation during post-transcriptional processing of RNA.

Chromatography, High Pressure Liquid↗
PubMed1987

Structure determination of a new fluorescent tricyclic nucleoside from archaebacterial tRNA.

A highly fluorescent nucleoside was detected in enzymatic digests of the extremely thermophilic archaebacterium Sulfolobus solfataricus by combined liquid chromatography-mass spectrometry (LC/MS). Following isolation, the structure was determined primarily by mass spectrometry, to be 3-(beta-D-ribofuranosyl)-4,9-dihydro-4,6,7-trimethyl-9-oxoimidazo[ 1, 2-a]purine (mimG), a new derivative of the Y (wye) nucleoside. The structural assignment was verified by comparison of the base released by acid hydrolysis with the corresponding synthetic base, using mass spectrometry, chromatography, and UV absorption and fluorescence properties. Nucleoside mimG was also detected by LC/MS in hydrolysates of the thermophiles Thermoproteus neutrophilus and Pyrodictium occultum. These results constitute the first finding of a member of the hypermodified Y family of nucleosides in archaebacteria.

Archaea↗
PubMed1986

Formation of 8-hydroxyguanine moiety in cellular DNA by agents producing oxygen radicals and evidence for its repair.

8-Hydroxydeoxyguanosine (8-OH-dG) was detected in DNA isolated from HeLa cells after the cells in tissue culture had been irradiated with X-rays and from the liver of mice after the whole animals had been irradiated with gamma-rays. The amounts of 8-OH-dG in DNA after in vivo irradiation were three orders of magnitude lower than those after in vitro irradiation (0.008-0.032 8-OH-dG residue/10(5) dG/krad). The 8-OH-dG produced in liver DNA by irradiation of mice decreased with time, suggesting the presence of a repair enzyme(s) acting on 8-OH-dG in mouse liver. Treatment of Salmonella typhimurium cells with hydrogen peroxide also caused increase in the 8-OH-dG content. These results indicate that 8-OH-dG is formed in vivo in cellular DNA on treatment with various oxygen radical-producing agents and that it is repairable.

8-Hydroxy-2'-Deoxyguanosine↗
PubMed1985

Photoaffinity labeling of diphtheria toxin fragment A with NAD: structure of the photoproduct at position 148.

Irradiation of mixtures of diphtheria toxin fragment A and [carbonyl-14C]NAD with UV light (253.7 nm) is known to induce efficient transfer of the radiolabel to position 148, corresponding to glutamic acid in the unmodified protein. Here we report the structure of the photoproduct at position 148, as determined by chemical and photochemical methods, fast-atom-bombardment mass spectrometry, and nuclear magnetic resonance. The photoproduct [an alpha-amino-gamma-(6-nicotin-amidyl)butyric acid residue] contains the entire nicotinamide moiety of NAD linked via its number 6 carbon to the decarboxylated gamma-methylene carbon of Glu-148. No portion of the ADP-ribosyl group of NAD is present. These findings are consistent with the idea that Glu-148 lies at or near the catalytic center of diphtheria toxin.

ADP Ribose Transferases↗
PubMed1983

Analysis of modified bases in DNA by stable isotope dilution gas chromatography-mass spectrometry: 5-methylcytosine.

A sensitive assay for 5-methylcytosine in DNA has been developed based on stable isotope dilution gas chromatography-mass spectrometry with selected ion monitoring. 5-[( 2H3]-Methyl)cytosine and [methyl-2H3]thymine have been synthesized as internal standards for analysis of DNA following acid digestion, conversion of pyrimidines to volatile t-butyldimethylsilyl derivatives, and separation in 3 min by gas chromatography. Submicrogram amounts of DNA have been analyzed for 5-methylcytosine content in the range 0.02-1.5 mol%. The estimated limit of quantitative measurement is 0.3 pmol of methylated base in a DNA hydrolysate. The method is compared with other techniques for quantitative measurement of methylated bases in DNA, and 5-methylcytosine levels and precision of analysis for calf thymus, pBR322, and phi X-174 DNAs are reported and compared with literature values. The method can readily be adapted to the accurate high-sensitivity analysis of other methylated bases in DNA.

5-Methylcytosine↗
PubMed1983

Purification and characterization of the unusual deoxynucleoside, alpha-N-(9-beta-D-2'-deoxyribofuranosylpurin-6-yl)glycinamide, specified by the phage Mu modification function.

Bacteriophage Mu encodes a protein that modifies approximately equal to 15% of DNA adenine residues to a new and unusual form. Modified DNA was enzymatically digested to deoxynucleosides, and the products were fractionated by HPLC. A modified adenine nucleoside, designated dA'x, was purified and its molecular structure was established by mass spectrometry. We show that dA'x is alpha-N-(9-beta-D-2'-deoxyribofuranosylpurin-6-yl)-glycinamide. The dA'x obtained from DNA was indistinguishable from the synthetic product with respect to its chromatographic behavior (HPLC and gas chromatography) and mass spectrum. Acid hydrolysis degrades dA'x to produce N6-carboxymethyladenine; this compound corresponds to the base Ax observed in earlier studies.

Chromatography, High Pressure Liquid↗
PubMed1982

Identification of nucleosides in hydrolysates of transfer RNA by high-resolution mass spectrometry.

High-resolution mass spectrometry has been investigated as a technique for identification of modified nucleosides in unfractionated hydrolysates of transfer RNA. The method is based on recognition of predetermined sets of exact mass values which are characteristic of individual nucleoside components. Mass spectra are photographically recorded in a non-time-resolved fashion, so that differing rates of nucleoside vaporization are of no consequence. Experimental parameters of sensitivity, spectrometer resolution and rates of vaporization were studied. Under typical conditions, 1-4 micrograms of tRNA are hydrolyzed, converted to volatile trimethylsilyl derivatives, and mass spectra of the resulting mixture recorded during 3 min at resolution 20 000; 75-100% of the minor nucleosides are usually identified in a single recording. The method is complementary to conventional methods of identification which rely on chromatographic mobilities, and can in principle be generally applied to recognition of biologically or chemically modified bases in nucleic acid.

Chemical Phenomena↗
PubMed1980

Structure of an amniotic fluid component, 7-(4,5-cis-dihydroxy-1-cyclopenten-3-ylaminomethyl)-7-deazaguanine (queuine), a substrate for tRNA: guanine transglycosylase.

A compound present in bovine amniotic fluid, and which is a substrate for tRNA:guanine transglycosylase, has been structurally characterized as queuine, a hypermodified base which occurs in transfer RNA. The structure assignment is based on mass spectrometry, proton nuclear magnetic resonance, and ultraviolet absorbance data.

Amniotic Fluid↗
PubMed1978

Origin of nitrogen in the biosynthesis of carbamoylpolyoxamic acid.

The biosynthetic origin of the carbamoylpolyoxamic sidechain of the polyoxin nucleoside peptide antibiotics was studied using 15N labeled alpha-amino-S-hydroxyvaleric acid as precursor. Gas chromatography mass spectrometry of polyoxamic acid isolated from culture filtrates of Streptomyces cacaoi var. asoensis showed incorporation of the nitrogen atom of alpha-amino-S-hydroxyvaleric acid into carbamoylpolyoxamic acid with an isotopic dilution factor of 23, thus establishing the intermediacy of alpha-amino-S-hydroxyvaleric acid and determination of the full biosynthetic pathway.

Amino Acids↗
PubMed1978

Characterization of N6, O2-dimethyladenosine from nuclear RNA of Novikoff hepatoma.

A modified nucleoside was isolated from low molecular weight nuclear RNA of Novikoff hepatoma, the nucleotide sequence of which was reported earlier. The structure of this novel nucleoside was shown to be N6, O2'-dimethyladenosine (m6Am) by mass spectrometry of its trimethylsilyl derivative and by comparison with the mass spectra of N6- and O-2' monmethyl model compounds. The complete characterization was carried out using 0.04 A260 units (2 microgram) of m6Am.

Adenosine↗
PubMed1978

Structure determination of a nucleoside Q precursor isolated from E. coli tRNA: 7-(aminomethyl)-7-deazaguanosine.

A precursor of modified nucleoside Q isolated from E. coli methyl-deficient tRNA was determined to be 7-(aminomethyl)-7-deazaguanosine. The structure was deduced by means of its chromatographic and electrophoretic mobilities, and UV and mass spectra, in addition to comparison with the synthesized authentic compound. The same molecule is also found in tRNA of an E. coli mutant selected for deficient synthesis of modified nucleosides.

Chromatography, Thin Layer↗
PubMed1975

Structure of the modified nucleoside Q isolated from Escherichia coli transfer ribonucleic acid. 7-(4,5-cis-Dihydroxy-1-cyclopenten-3-ylaminomethyl)-7-deazaguanosine.

The structure of the unknown modified nucleoside Q, which is present in the first position of the anticodons of Escherichia coli tRNA Tyr, tRNA His, tRNA Asn, tRNA Asp, is proposed to be 7-(4,5-cis-dihydroxy-1-cyclopenten-3-ylaminomethyl)-7-deazaguanosine (1). The structure of Q was deduced by means of its uv absorption, mass spectrometry, proton magnetic resonance spectroscopy, and studies of its chemical reactivity. The structure of Q is unique since it is a derivative of 7-deazaguanosine having cyclopentenediol in the side chain at the C-7 position. This is the first example of purine skeleton modification in a nucleoside from tRNA.

Anticodon↗