1887

Abstract

6-Hydroxy--nicotine oxidase (6-HDNO) of , an enzyme inducible by -nicotine, contains FAD covalently bound via an 8a-N(3)His linkage. Expression of the gene encoding 6-HDNO and flavinylation of the protein were studied in . In this heterologous system the following findings were made. 1. An enzymically active covalently flavinylated 6-HDNO of normal size can be expressed in . 2. The natural promoter of the 6-HDNO gene appeared inefficient in . The promoter, when inserted upstream of the promoter, increased 6-HDNO expression >50-fold. 3. Expression of the 6-HDNO gene from plasmids in was, independently of the promoter construct used, stimulated more than fivefold by -nicotine in the growth medium. It is concluded that flavinylation of 6-HDNO is possibly autocatalytic and mediated by factors generally found in bacterial cells.

Loading

Article metrics loading...

/content/journal/micro/10.1099/00221287-135-5-1093
1989-05-01
2024-04-27
Loading full text...

Full text loading...

/deliver/fulltext/micro/135/5/mic-135-5-1093.html?itemId=/content/journal/micro/10.1099/00221287-135-5-1093&mimeType=html&fmt=ahah

References

  1. Arwert F., Venema G. 1973; Transformation in Bacillus subtilis. Fate of newly introduced transforming DNA. Molecular and General Genetics 123:185–198
    [Google Scholar]
  2. Berger S.L., Kimmel A.R. 1987; Guide to molecular cloning techniques. Methods in Enzymology 152:33–170
    [Google Scholar]
  3. Boyer H.W., Roulland-Dussoix D. 1969; A complementation analysis of the restriction and modification of DNA in Escherichia coli . Journal of Molecular Biology 41:459–472
    [Google Scholar]
  4. Brandsch R., Bichler V. 1985; In vivo and in vitro expression of the 6-hydroxy-d-nicotine oxidase gene of Arthrobacter oxidans, cloned into Escherichia coli, as an enzymatically active, covalently flavinylated polypeptide. FEBS Letters 192:204–208
    [Google Scholar]
  5. Brandsch R., Bichler V. 1986; Studies in vitro on the flavinylation of 6-hydroxy-d-nicotine oxidase. European Journal of Biochemistry 160:285–289
    [Google Scholar]
  6. Brandsch R., Bichler V. 1987; Covalent flavinylation of 6-hydroxy-d-nicotine oxidase involves an energy-requiring process. FEBS Letters 224:121–124
    [Google Scholar]
  7. Brandsch R., Faller W., Schneider K. 1986; Plasmid pAOl of Arthrobacter oxidans encodes 6- hydroxy-d-nicotine oxidase: cloning and expression of the gene in Escherichia coli . Molecular and General Genetics 202:96–101
    [Google Scholar]
  8. Brandsch R., Bichler V., Nagursky H. 1987a; Covalent flavinylation of 6-hydroxy-d-nicotine oxidase analyzed by partial deletions of the gene. European Journal of Biochemistry 165:559–564
    [Google Scholar]
  9. Brandsch R., Hinkkanen A.E., Mauch L., Nagursky H., Decker K. 1987b; 6-Hydroxy-d-nicotine oxidase of Arthrobacter oxidans. Gene structure of the flavoenzyme and its relationship to 6-hydroxy-l-nicotine oxidase. European Journal of Biochemistry 167:315–320
    [Google Scholar]
  10. Brandsch R., Fiinkkanen A.E., Mauch L., Nagursky H., Decker K. 1987c; The flavin binding sites of 6-hydroxy-d- and 6-hydroxy-l-nicotine oxidases. In Flavins and Flavoproteins pp. 747–750 Edmondson D.E., Mccormick D.B. Edited by New York: Walter de Gruyter;
    [Google Scholar]
  11. Brühmüller M., Möhler H., Decker K. 1972; Covalently bound flavin in 6-d-hydroxynicotine oxidase from Arthrobacter oxidans. Purification and properties of 6-d-hydroxynicotine oxidase. European Journal of Biochemistry 29:143–151
    [Google Scholar]
  12. Cecchini G., Perl M., Lipsick J., Singer T.P., Kearney E.B. 1979; Transport and binding of riboflavin by Bacillus subtilis . Journal of Biological Chemistry 254:7295–7301
    [Google Scholar]
  13. Cecchini G., Kearney E.B., Jones H., Gunsalus R.P. 1985; Studies on the attachment of covalent flavins to fumarate reductase from Escherichia coli . Federation Proceedings 44:1766
    [Google Scholar]
  14. Cecchini G., Ackrell B.A.C., Whittiker K., Blout M., Gunsalus R.P. 1988; Fumarate reductase containing non-covalently bound FAD. FASEB Journal 2:354
    [Google Scholar]
  15. Cole S.T. 1982; Nucleotide sequence coding for the flavoprotein subunit of the fumarate reductase of Escherichia coli . European Journal of Biochemistry 122:479–484
    [Google Scholar]
  16. Decker K. 1982; Biosynthesis of covalent flavoproteins. In Flavins and Flavoproteins pp. 465–472 Massey V., Williams C.H. Edited by Amsterdam: Elsevier North Holland;
    [Google Scholar]
  17. Decker K., Bleeg H. 1965; Induction and purification of stereospecific nicotine oxidizing enzymes. Biochimica et biophysica acta 105:313–334
    [Google Scholar]
  18. Decker K., Gries F.A., Brühmüller M. 1961; Über den Abbau des Nicotins durch Bakterien- enzyme. III. Stoffwechselstudien an Zellfreien Ex- trakten. Hoppe-Seyler𠆖s Zeitschrift für Physiologische Chemie 323:249–263
    [Google Scholar]
  19. Doi R.H., Wang L.-F. 1986; Multiple procaryotic ribonucleic acid polymerase sigma factors. Microbiological Reviews 50:227–243
    [Google Scholar]
  20. Eberwein H., Gries F.A., Decker K. 1961; Über den Abbau des Nicotins durch Bakterien- enzyme. II. Isolierung und Charakteriserung eines nicotin-abbauenden Bodenbakteriums. Hoppe Seyler’s Zeitschrift für Physiologische Chemie 323:236–248
    [Google Scholar]
  21. Gloger M., Decker K. 1969; Zum Mechanismus der Inducktion nicotinabbauender Enzyme in Arth-robacter oxidans . Zeitschrift für Naturforschung 24b:1016–1025
    [Google Scholar]
  22. Gryczan T.J., Contente S., Dubnau D. 1978; Characterization of Staphylococcus aureus plasmids introduced by transformation into Bacillus subtilis . Journal of Bacteriology 134:318–323
    [Google Scholar]
  23. Hager P.W., Rabinowitz J.C. 1985; Translational specificity in Bacillus subtilis . In The Molecular Biology of the Bacilli 2 pp. 1–32 Dubnau D.A. Edited by New York: Academic Press;
    [Google Scholar]
  24. Hederstedt L. 1983; Succinate dehydrogenase mutants of Bacillus subtilis lacking covalently bound flavin in the flavoprotein subunit. European Journal of Biochemistry 132:589–593
    [Google Scholar]
  25. Hederstedt L., Bergman T., Jörnvall H. 1987; Processing of Bacillus subtilis succinate dehydrogenase and cytochrome 6558 polypeptides. Lack of covalently bound flavin in the Bacillus enzyme expressed in Escherichia coli . FEBS Letters 213:385–390
    [Google Scholar]
  26. Ish-Horowicz D., Burke J.F. 1981; Rapid and efficient cosmid cloning. Nucleic Acids Research 9:2989–2998
    [Google Scholar]
  27. Laemmli U.K. 1970; Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature; London: 227680–685
    [Google Scholar]
  28. Maguire J.J., Magnusson K., Hederstedt L. 1986; Bacillus subtilis mutant succinate dehydrogenase lacking covalently bound flavin: identification of the primary defect and studies on the iron- sulfur clusters in mutated and wildtype enzyme. Biochemistry 25:5202–5208
    [Google Scholar]
  29. Meun L., Magnusson K., Rutberg L. 1987; Identification of the promoter of the Bacillus subtilis sdh operon. Journal of Bacteriology 169:3232–3236
    [Google Scholar]
  30. Möhler H., Brühmüller M., Decker K. 1972; Covalently bound flavin in d-6-hydroxynicotine oxidase. Identification of the 8aα(N-3-histidyl)-ribo- flavin linkage between FAD and the apoenzyme. European Journal of Biochemistry 29:152–155
    [Google Scholar]
  31. Nagursky H., Bichler V., Brandsch R. 1988; Phosphoenolpyruvate-dependent flavinylation of 6-hydroxy-d-nicotine oxidase. European Journal of Biochemistry 177:319–325
    [Google Scholar]
  32. Phillips M.K., Hederstedt L., Hasnain S., Rutberg L., Guest J.R. 1987; Nucleotide sequence encoding the flavoprotein and iron-sulfur protein subunits of the Bacillus subtilis PY79 succinate dehydrogenase complex. Journal of Bacteriology 169:864–873
    [Google Scholar]
  33. Salach J., Walker W.H., Singer T.P., Ehrenberg A., Hemmerich P., Ghisla S., Hartmann U. 1972; Studies on succinate dehydrogenase. Site of attachment of the covalently bound flavin to the peptide chain. European Journal of Biochemistry 26:267–278
    [Google Scholar]
  34. Shaw W.V. 1975; Chloramphenicol acetyltrans- ferase from chloramphenicol resistant bacteria. Methods in Enzymology 43:737–755
    [Google Scholar]
  35. Singer T.P., McIntire W.S. 1984; Covalent attachment of flavin to flavoproteins: occurrence, assay and synthesis. Methods in Enzymology 106:369–378
    [Google Scholar]
  36. Timoco I. JR Borer P.N., Dengler B., Levine M.D., Uhlenbeck O.C., Crothers D.M., Gralla J. 1973; Improved estimation of secondary structure in ribonucleic acids. Nature; London: 24640–41
    [Google Scholar]
  37. Williams D.M., Duvall E.J., Lovett P.S. 1981; Cloning restriction fragments that promote expression of a gene in Bacillus subtilis . Journal of Bacteriology 146:1162–1165
    [Google Scholar]
  38. Woese C.R. 1987; Bacterial evolution. Microbiological Reviews 51:221–271
    [Google Scholar]
  39. Wood D., Darlison M.G., Wilde R.J., Guest J.R. 1984; Nucleotide sequence encoding the flavoprotein and hydrophobic subunits of the succinate dehydrogenase of Escherichia coli . Biochemical Journal 222:519–534
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/micro/10.1099/00221287-135-5-1093
Loading
/content/journal/micro/10.1099/00221287-135-5-1093
Loading

Data & Media loading...

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error