Crystal and molecular structures of 2-amino-3-methylimidazo-[4,5-f]guinoline, a novel potent mutagen found in broiled food.
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Biomedical subjects
Publications and source records attributed to S Nishimura.
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Several DNA fragments carrying tRNA genes have been cloned from EcoRI endonuclease digests of Escherichia coli DNA. Using cloned DNA, the sequence of the region around the distal gene for tRNA1Asp (F(or G)) in the E. coli ribosomal RNA operon [rrnF(or G)] has been determined. In the distal portion of rrnF(or G), the genes for 23S, 5S rRNA and tRNA1Asp (F(or G)) are located in that order and separated by intergenic spacers of 93 and 52 base pairs, respectively. A possible hairpin structure, with its center between the 22nd and 23rd base pair downstream from the 3'-end of the tRNA1Asp(F(or G)) gene, followed by a sequence of eight thymidine residues was identified as the transcription termination signal for rrnF(or G). The termination is rho-independent, at least in vitro, and occurs within the region of the contiguous thymidine residues. A possible promoter for a protein gene is present about 50 base pairs downstream from the rrnF(or G) terminator.
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The nucleotide sequence of an E. coli isoleucine tRNA (tRNAIle minor) specific for the codon AUA was determined by postlabeling procedures using only 2.5 micrograms (0.05 A260 unit) of the material. The sequence was pG-G-C-C-C-C-U-s4U-A-G-C-U-C-A-G-U-Gm-G-D-D-A-G-A-G-C-A-A-G-C-G-A-C-U-N+-A-U-t6A-A-psi-C-G-C-U-U-G-m7G-acp3U-C-G-C-U-G-G-T-psi-C-A-A-G-U-C-C-A-G-C-A-G-G-G-G-C-C-A-C-C-AOH. The nucleotide sequences in the regions of the D arm and T psi C arm of tRNAIle minor were quite similar to the corresponding regions of tRNAIle major. However, the sequences in the CCA stem and anticodon stem of tRNAIle minor were different from those of tRNAIle major. The overall homology between the two isoleucine tRNAs was 68%. E. coli tRNALys, tRNAMet, tRNAValIIA and tRNAArg also have relatively high sequence homology with tRNAIle minor.
The primary structure of Escherichia coli tRNA UUR Le which recognizes the UU series of codons has been determined. The sequence is pG-C-C-C-G-G-A-s4U-G-G-U-G-G-A-A-D-C-Gm-C-D-A-G-A-C-A-C-A-A-G-G-G-A-psi-U-N-A-A-ms2i6A-A-psi-C-C-C-C-U-C-G-G-C-G-G-C-G-U-U-C-G-C-G-C-U-G-U-G-C-G-G-G-T-psi-C-A-A-G-U-C-C-C-G-C-U-C-C--G-G-G-U-A-C-C-A. The chain length of tRNA UUR Leu is 87 residues, the same as other E. coli tRNA Leu s and T4 phage-coded tRNA Leus. Its sequence is especially similar to that of E. coli tRNA2 Leu in the Darm and T psi C arm regions. E. coli tRNA UUR Leu contains an unknown modified nucleoside in the first position of the anticodon and was shown by mass spectrometry and chemical degradation to be an adenosine derivativee. Addition of tRNA UUR Leu to a cell-free protein-synthesizing system with high Mg2+ concentration resulted in the formation of polyleucine miscoded by poly(U), indicating that the unknown modified nucleoside exhibits a tendency to recognize U under certain conditions.
Transfer ribonucleic acid (tRNA) guanine transglycosylase (guanine insertion enzyme) was isolated from rat liver and extensively purified. The enzyme catalyzes an exchange of queuine (the base of queuosine, Q) as well as its precursors and guanine for guanine originally located in the first position of the anticodon of "undermodified" tRNATyr, tRNAHis, tRNAAsn, and tRNAAsp from an Escherichia coli mutant or rat ascites hepatoma cells. This is in contrast to the previous observation that E. coli tRNA-guanine transglycosylase catalyzes the exchange of queuine precursors, such as 7-(aminoethyl)-7-deazaguanine and 7-cyano-7-deazaguanine, but not of queuine itself [Okada, N., Noguchi, S. Kasai, H., Shindo-Okada, N., Ohgi, T., Goto, T., & Nishimura, S. (1979) J. Biol. Chem. 254, 3067-3073]. The Km value for queuine of the rat liver enzyme is 9.2 X 10(-7) M, much lower than the values for the bases of queuosine precursors or guanine. Thus, the actual substrate for tRNA-guanine transglycosylase in queuosine biosynthesis in vivo in rat liver may not be 7-(aminomethyl)-7-deazaguanine, which is thought to be an actual substrate guanine, the E. coli system. Queuine or some queuine derivative may be the actual substrate for the tRNA-guanine transglycosylase reaction in the biosynthesis of Q in tRNA of mammalian cells. 6-Thioguanine and 8-azaguanine are also found to be good substrates.
An experimental study using adult dog was conducted on the impression of the colon whereby a screening test is done by making a mold of the intestine with sodium alginate. The mold of the intestine could be obtained easily and safely in a short time. Protrusions about 2 mm in diameter corresponding to lymphoid tissue of the rectum and sigmoid colon and shallow depressions on the surface of the protrusions were exactly imprinted on the surface of the mild. Compared with the barium study and endoscopy, this method requires neither special equipment nor high-grade technique, can be done easily in a short time and was not inferior in the diagnosis accuracy. Sodium alginate used in this method is a safe substance which is being used clinically as a plasma expander and laxative in addition to being used as an additive to various foods. When injected into the intestine, it exerted no injurious action whatsoever. from these findings, the possibility of this method being used clinically was considered high.
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The major mutagenic component of fried beef has been isolated using a series of chromatographic steps. The pure compound has been analyzed by low and high resolution mass spectroscopy and nuclear magnetic resonance spectroscopy. The results indicate that the molecular weight of this extremely mutagenic compound is 198, with an elemental composition of C11H10N4. The compound is different from the known mutagenic pyrolysis products of amino acids or proteins.
The potent mutagens Trp-P-1 (3-amino-1, 4-dimethyl-5H-pyrido-[4,3-b]-indole) and Trp-P-2 (3-amino-1-methyl-5H-pyrido[4,3-b]indole) are known to be produced by pyrolysis of tryptophan [8]. To determine whether such mutagens are produced by cooking foods, the fractions obtained from broiled sardines cooked in the ordinary way were analysed by gas chromatography/mass spectrometry. The results showed that 13.3 ng of Trp-P-1 and 13.1 ng of Trp-P-2 were, in fact, present per gram of broiled sardines.
Three species of methionine tRNAs and phenylalanine, tyrosine, and isoleucine tRNAs were purified from an extreme thermophile, Thermus thermophilus HB8. Formylation studies of the three methionine tRNAs and their codon-specific binding activities to ribosomes showed that two of them (named tRNAf1Met and tRNAf2Met) were initiator tRNAs and the other (named tRNAmMet) was a non-initiator. The tRNAs from T. thermophilus all had melting temperatures of up to ten degrees higher than the corresponding species from E. coli. Most of the species also had slightly higher G+C contents than the corresponding species of E. coli, and each of them contained one mol each of the modified nucleosides, O2'-methylguanosine (Gm), 2-thioribothymidine (s2T), and 1-methyladenosine (m1A). Their high melting temperatures could be explained by their high G+C contents and the presence of the modified nucleosides, espically s2T. Comparison of the melting temperatures of T. thermophilus tRNAf2Met with those of E. coli tRNAfMet and tRNAmMet at different magnesium concentrations showed that magnesium was also a factor in the thermostability of the thermophile tRNA.
Discadenine,3-(3-amino-3-carboxypropyl)-N6-delta 2-isopentenyladenine, which inhibits spore germination, was previously found in Dictyostelium discoideum. Studies on the distribution of discadenine in different species of cellular slime molds by high-pressure liquid chromatography showed that discadenine is present in D. discoideum, Dictyostelium purpureum, and Dictyostelium mucoroides, but not in Dictyostelium minutum, Polysphondylium violaceum, or Polysphondylium pallidum. Discadenine synthetase, which is involved in biosynthesis of discadenine with N6-delta 2-isopentenyladenine as substrate, was only detected in cells of the former three species. In addition, discadenine inhibited spore germination only in these three species. These results clearly demonstrate that discadenine is produced as an inhibitor of spore germination in the species of cellular slime molds in which the acrasin is cyclic adenosine 5'-monophosphate (AMP). This means that there is a structural and biochemical correlation between the spore germination inhibitor and the acrasin, since 5'-AMP, a direct precursor in discadenine biosynthesis, can be derived from cyclic AMP by hydrolysis with cyclic AMP phosphodiesterase.
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Relation of the intimal change in aging of various arteries to progression of atherosclerosis and the morphology of regression of atherosclerotic lesion were discussed. The course of regression of experimental atherosclerosis in rabbits and its histological findings were described.
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