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. 1974 Nov;143(2):285–294. doi: 10.1042/bj1430285

Studies on chemical modification of thionucleosides in the transfer ribonucleic acid of Escherichia coli

Yarlagadda S Prasada Rao 1, Joseph D Cherayil 1
PMCID: PMC1168383  PMID: 4376941

Abstract

35S-labelled tRNA from Escherichia coli was treated with chemical reagents such as CNBr, H2O2, NH2OH, I2, HNO2, KMnO4 and NaIO4, under mild conditions where the four major bases were not affected. Gel filtration of the treated tRNA showed desulphurization to various extents, depending on the nature of the reagent. The treated samples after conversion into nucleosides were chromatographed on a phosphocellulose column. NH2OH, I2 and NaIO4 reacted with all the four thionucleosides of E. coli tRNA, 4-thiouridine (s4U), 5-methylaminomethyl-2-thiouridine (mnm5s2U), 2-thiocytidine (s2C) and 2-methylthio-N6-isopentenyladenosine (ms2i6A), to various extents. CNBr, HNO2 and NaHSO3 reacted with s4U, mnm5s2U and s2C, but not with ms2i6A. KMnO4 and H2O2 were also found to react extensively with thionucleosides in tRNA. Iodine oxidation of 35S-labelled tRNA showed that only 6% of the sulphur was involved in disulphide formation. Desulphurization of E. coli tRNA with CNBr resulted in marked loss of acceptor activities for glutamic acid, glutamine and lysine. Acceptor activities for alanine, arginine, glycine, isoleucine, methionine, phenylalanine, serine, tyrosine and valine were also affected, but to a lesser extent. Five other amino acids tested were almost unaffected. These results indicate the fate of thionucleosides in tRNA when subjected to various chemical reactions and the involvement of sulphur in aminoacyl-tRNA synthetase recognition of some tRNA species of E. coli.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Burrows W. J., Armstrong D. J., Skoog F., Hecht S. M., Boyle J. T., Leonard N. J., Occolowitz J. Cytokinin from soluble RNA of Escherichia coli: 6-(3-methyl-2-butenylamino)-2-methylthio-9-beta-D-ribofuranosylpurine. Science. 1968 Aug 16;161(3842):691–693. doi: 10.1126/science.161.3842.691. [DOI] [PubMed] [Google Scholar]
  2. Carbon J. A., Hung L., Jones D. S. A reversible oxidative in activation of specific transfer RNA species. Proc Natl Acad Sci U S A. 1965 May;53(5):979–986. doi: 10.1073/pnas.53.5.979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Carbon J., Curry J. B. Genetically and chemically derived missense suppressor transfer RNA's with altered enzymic aminoacylation rates. J Mol Biol. 1968 Dec 14;38(2):201–216. doi: 10.1016/0022-2836(68)90406-3. [DOI] [PubMed] [Google Scholar]
  4. Carbon J., David H., Studier M. H. Thiobases in Escherchia coli Transfer RNA: 2-Thiocytosine and 5-Methylaminomethyl-2-thiouracil. Science. 1968 Sep 13;161(3846):1146–1147. doi: 10.1126/science.161.3846.1146. [DOI] [PubMed] [Google Scholar]
  5. Carbon J., David H. Studies on the thionucleotides in transfer ribonucleic acid. Addition of N-ethylmaleimide and formation of mixed disulfides with thiol compounds. Biochemistry. 1968 Nov;7(11):3851–3858. doi: 10.1021/bi00851a010. [DOI] [PubMed] [Google Scholar]
  6. Chambers R. W. On the recognition of tRNA by its aminoacyl-tRNA ligase. Prog Nucleic Acid Res Mol Biol. 1971;11:489–525. doi: 10.1016/s0079-6603(08)60336-0. [DOI] [PubMed] [Google Scholar]
  7. Dirheimer G., Ebel J. P., Bonnet J., Gangloff J., Keith G., Krebs B., Kuntzel B., Roy A., Weissenbach J., Werner C. Structure primaire des tRN. Biochimie. 1972;54(2):127–144. doi: 10.1016/s0300-9084(72)80097-x. [DOI] [PubMed] [Google Scholar]
  8. Doctor B. P., Wayman B. J., Cory S., Rudland P. S., Clark B. F. Studies on the Escherichia coli Methionine transfer ribonucleic acids. Eur J Biochem. 1969 Mar;8(1):93–100. doi: 10.1111/j.1432-1033.1969.tb00500.x. [DOI] [PubMed] [Google Scholar]
  9. Faulkner R. D., Uziel M. Iodine modification of E. coli tRNA Phe : reversible modification of 2-methylthio-N 6 -isopentenyladenosine and lack of disulfide formation. Biochim Biophys Acta. 1971 May 27;238(3):464–474. doi: 10.1016/0005-2787(71)90620-4. [DOI] [PubMed] [Google Scholar]
  10. Furuichi Y., Wataya Y., Hayatsu H., Ukita T. Chemical modification of tRNA-Tyr-yeast with bisulfite. A new method to modify isopentenyladenosine residue. Biochem Biophys Res Commun. 1970 Dec 9;41(5):1185–1191. doi: 10.1016/0006-291x(70)90211-1. [DOI] [PubMed] [Google Scholar]
  11. Goodman H. M., Abelson J., Landy A., Brenner S., Smith J. D. Amber suppression: a nucleotide change in the anticodon of a tyrosine transfer RNA. Nature. 1968 Mar 16;217(5133):1019–1024. doi: 10.1038/2171019a0. [DOI] [PubMed] [Google Scholar]
  12. Gross H. J., Czerny E. M. Temperature-dependent inactivation of tRNA Tyr E.coli acceptor function with iodine: influence of the 3'-terminal pCpA sequence. FEBS Lett. 1973 Mar 15;30(3):347–350. doi: 10.1016/0014-5793(73)80685-4. [DOI] [PubMed] [Google Scholar]
  13. Hall R. H. N6-(delta 2-isopentenyl)adenosine: chemical reactions, biosynthesis, metabolism, and significance to the structure and function of tRNA. Prog Nucleic Acid Res Mol Biol. 1970;10:57–86. doi: 10.1016/s0079-6603(08)60561-9. [DOI] [PubMed] [Google Scholar]
  14. Hampel A., Cherayil J., Bock R. M. Specific aggregation of yeast glycine tRNA. Biochim Biophys Acta. 1971 Jan 28;228(2):482–491. doi: 10.1016/0005-2787(71)90053-0. [DOI] [PubMed] [Google Scholar]
  15. Harris C. L., Titchener E. B., Cline A. L. Sulfur-deficient transfer ribonucleic acid in a cysteine-requiring, "relaxed" mutant of Escherichia coli. J Bacteriol. 1969 Dec;100(3):1322–1327. doi: 10.1128/jb.100.3.1322-1327.1969. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Hayatsu H., Inoue M. The oxygen-mediated reaction between 4-thiouracil derivatives and bi- sulfite. Isolation and characterization of 1-methyluracil 4-thiosulfate as an intermediate in the formation of 1-methyluracil-4-sulfonate. J Am Chem Soc. 1971 May 5;93(9):2301–2306. doi: 10.1021/ja00738a033. [DOI] [PubMed] [Google Scholar]
  17. Hayatsu H., Wataya Y., Kai K., Iida S. Reaction of sodium bisulfite with uracil, cytosine, and their derivatives. Biochemistry. 1970 Jul 7;9(14):2858–2865. doi: 10.1021/bi00816a016. [DOI] [PubMed] [Google Scholar]
  18. Hayatsu H., Wataya Y., Kazushige K. The addition of sodium bisulfite to uracil and to cytosine. J Am Chem Soc. 1970 Feb 11;92(3):724–726. doi: 10.1021/ja00706a062. [DOI] [PubMed] [Google Scholar]
  19. Hayward R. S., Weiss S. B. RNA thiolase: the enzymatic transfer of sulfur from cysteine to sRNA in Escherichia coli extracts. Proc Natl Acad Sci U S A. 1966 May;55(5):1161–1168. doi: 10.1073/pnas.55.5.1161. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Hecht S. M., Kirkegaard L. H., Bock R. M. Chemical modifications of transfer RNA species. Desulfurization with Raney nickel. Proc Natl Acad Sci U S A. 1971 Jan;68(1):48–51. doi: 10.1073/pnas.68.1.48. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Iida S., Chung K. C., Hayatsu H. The reaction of hydroxylamine with 4-thiouridine. Biochim Biophys Acta. 1973 May 10;308(2):198–204. doi: 10.1016/0005-2787(73)90149-4. [DOI] [PubMed] [Google Scholar]
  22. Iida S., Hayatsu H. Chemical transformation of 4-thiouridine with nitrous acid. Biochem Biophys Res Commun. 1971 Apr 2;43(1):163–167. doi: 10.1016/s0006-291x(71)80101-8. [DOI] [PubMed] [Google Scholar]
  23. Imura N., Schwam H., Chambers R. W. Transfer RNA. 3. Reconstitution of alanine acceptor activity from fragments produced by specific cleavage of tRNA Ala-II at its anticodon. Proc Natl Acad Sci U S A. 1969 Apr;62(4):1203–1209. doi: 10.1073/pnas.62.4.1203. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Kochetkov N. K., Budowsky E. I. The chemical modification of nucleic acids. Prog Nucleic Acid Res Mol Biol. 1969;9:403–438. doi: 10.1016/s0079-6603(08)60773-4. [DOI] [PubMed] [Google Scholar]
  25. Kućan Z., Freude K. A., Kućan I., Chambers R. W. Aminoacylation of bisulphite-modified yeast tyrosine transfer RNA. Nat New Biol. 1971 Aug 11;232(2):177–179. doi: 10.1038/newbio232177a0. [DOI] [PubMed] [Google Scholar]
  26. Lipsett M. N., Doctor B. P. Studies on tyrosine transfer ribonucleic acid, a sulfur-rich species from Escherichia coli. J Biol Chem. 1967 Sep 25;242(18):4072–4077. [PubMed] [Google Scholar]
  27. Lipsett M. N., Peterkofsky A. Enzymatic thiolation of E. coli sRNA. Proc Natl Acad Sci U S A. 1966 May;55(5):1169–1174. doi: 10.1073/pnas.55.5.1169. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Lipsett M. N. The isolation of 4-thiouridylate disulfide from oxidized transfer ribonucleic acid of Escherichia coli. J Biol Chem. 1967 Sep 25;242(18):4067–4071. [PubMed] [Google Scholar]
  29. Lipsett M. N. The isolation of 4-thiouridylic acid from the soluble ribonucleic acid of Escherichia coli. J Biol Chem. 1965 Oct;240(10):3975–3978. [PubMed] [Google Scholar]
  30. Mirzabekov A. D., Lastity D., Levina E. S., Bayev A. A. Localization of two recognition sites in yeast valine tRNA I. Nat New Biol. 1971 Jan 6;229(1):21–22. doi: 10.1038/newbio229021a0. [DOI] [PubMed] [Google Scholar]
  31. Miura K. Specificity in the structure of transfer RNA. Prog Nucleic Acid Res Mol Biol. 1967;6:39–82. doi: 10.1016/s0079-6603(08)60524-3. [DOI] [PubMed] [Google Scholar]
  32. Nishimura S. Minor components in transfer RNA: their characterization, location, and function. Prog Nucleic Acid Res Mol Biol. 1972;12:49–85. [PubMed] [Google Scholar]
  33. Rao Y. S., Cherayil J. D. Separation of 35 S-labeled thionucleosides of Escherichia coli and Pseudomonas aeruginosa transfer RNAs on a phosphocellulose column. Biochim Biophys Acta. 1973 Feb 23;299(1):1–7. doi: 10.1016/0005-2787(73)90391-2. [DOI] [PubMed] [Google Scholar]
  34. Saneyoshi M., Anami T., Nishimura S., Samejima T. The change in optical activity of amino acid-specific Escherichia coli transfer RNA containing 4-thiouridylate by chemical modifications. Arch Biochem Biophys. 1972 Oct;152(2):677–684. doi: 10.1016/0003-9861(72)90264-0. [DOI] [PubMed] [Google Scholar]
  35. Saneyoshi M., Nishimura S. Alteration of codon recognition of Escherichia coli transfer RNA by modification with cyanogen bromide. Biochim Biophys Acta. 1967 Aug 22;145(1):208–210. doi: 10.1016/0005-2787(67)90680-6. [DOI] [PubMed] [Google Scholar]
  36. Saneyoshi M., Nishimura S. Selective inactivation of amino acid acceptor and ribosome-binding activities of Escherichia coli tRNA by modification with cyanogen bromide. Biochim Biophys Acta. 1971 Aug 12;246(1):123–131. doi: 10.1016/0005-2787(71)90077-3. [DOI] [PubMed] [Google Scholar]
  37. Saneyoshi M., Nishimura S. Selective modification of 4-thiouridylate residue in Escherichia coli transfer RNA with cyanogen bromide. Biochim Biophys Acta. 1970 Apr 15;204(2):389–399. doi: 10.1016/0005-2787(70)90158-9. [DOI] [PubMed] [Google Scholar]
  38. Scheit K. H. Uber die Synthese und Eigenschaften von 4-Thiouridylyl-(3'-5')-4-thiouridin, 4-Thiouridylyl-(3'-5')-uridin und Uridylyl-(3'-5')-4-thiouridin. Biochim Biophys Acta. 1968 Sep 24;166(2):285–293. [PubMed] [Google Scholar]
  39. Shugart L. Effect of selective chemical modification of 4-thiouridine of phenylalanine transfer ribonucleic acid on enzyme recognition. Arch Biochem Biophys. 1972 Feb;148(2):488–495. doi: 10.1016/0003-9861(72)90167-1. [DOI] [PubMed] [Google Scholar]
  40. Singhal R. P. Modification of Escherichia coli glutamate transfer ribonucleic acid with bisulfite. J Biol Chem. 1971 Sep 25;246(18):5848–5851. [PubMed] [Google Scholar]
  41. Sundharadas G., Katze J. R., Söll D., Konigsberg W., Lengyel P. On the recognition of serine transfer RNA's specific for unrelated codons by the same seryl-transfer RNA synthetase. Proc Natl Acad Sci U S A. 1968 Oct;61(2):693–700. doi: 10.1073/pnas.61.2.693. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Thiebe R., Zachau H. G. The role of the anticodon region in homologous and heterologous charging of tRNA-Phe. Biochem Biophys Res Commun. 1968 Oct 24;33(2):260–265. doi: 10.1016/0006-291x(68)90778-x. [DOI] [PubMed] [Google Scholar]
  43. Uziel M. Stability of 4,4'-di(thiouridine) and 4-thiouridine in alkali. Biochem Biophys Res Commun. 1966 Oct 5;25(1):105–108. doi: 10.1016/0006-291x(66)90646-2. [DOI] [PubMed] [Google Scholar]
  44. WHITFELD P. R., MARKHAM R. Natural configuration of the purine nucleotides in ribonucleic acids; chemical hydrolysis of the dinucleoside phosphates. Nature. 1953 Jun 27;171(4365):1151–1152. doi: 10.1038/1711151a0. [DOI] [PubMed] [Google Scholar]
  45. Walker R. T., RajBhandary U. L. E. coli tyrosine transfer RNA: chemical modification of thiouridine to uridine. Biochem Biophys Res Commun. 1970 Mar 12;38(5):907–914. doi: 10.1016/0006-291x(70)90807-7. [DOI] [PubMed] [Google Scholar]
  46. Walker R. T., RajBhandary U. L. Studies on polynucleotides. CI. Escherichia coli tyrosine and formylmethionine transfer ribonucleic acids: effect of chemical modification of 4-thiouridine to uridine on their biological properties. J Biol Chem. 1972 Aug 10;247(15):4879–4892. [PubMed] [Google Scholar]
  47. Yaniv M., Favre A., Barrell B. G. Structure of transfer RNA. Evidence for interaction between two non-adjacent nucleotide residues in tRNA from Escherichia coli. Nature. 1969 Sep 27;223(5213):1331–1333. doi: 10.1038/2231331a0. [DOI] [PubMed] [Google Scholar]
  48. Yano M., Hayatsu H. Permanganate oxidation of 4-thiouracil derivatives. Isolation and properties of I-substituted 2-pyrimidone 4-sulfonates. Biochim Biophys Acta. 1970 Feb 18;199(2):303–315. [PubMed] [Google Scholar]
  49. Yoshida M., Furuichi Y., Kaziro Y., Ukita T. The modification of nucleosides and nucleotides. IX. Inactivation of coding response of yeast tRNA containing inosine residue by cyanoethylation. Biochim Biophys Acta. 1968 Oct 29;166(3):636–645. [PubMed] [Google Scholar]
  50. Ziff E. B., Fresco J. R. Chemical transformation of 4-thiouracil nucleosides to uracil and cytosine counterparts. J Am Chem Soc. 1968 Dec 18;90(26):7338–7342. doi: 10.1021/ja01028a027. [DOI] [PubMed] [Google Scholar]

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