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Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1995 Jan 31;92(3):714–718. doi: 10.1073/pnas.92.3.714

Occurrence of enzymes involved in biosynthesis of indole-3-acetic acid from indole-3-acetonitrile in plant-associated bacteria, Agrobacterium and Rhizobium.

M Kobayashi 1, T Suzuki 1, T Fujita 1, M Masuda 1, S Shimizu 1
PMCID: PMC42690  PMID: 11607511

Abstract

The occurrence of a hitherto unknown pathway involving the action of two enzymes, a nitrile hydratase and an amidase for the biosynthesis of indole-3-acetic acid was discovered in phytopathogenic bacteria Agrobacterium tumefaciens and in leguminous bacteria Rhizobium. The nitrile hydratase acting on indole-3-acetonitrile was purified to homogeneity through only two steps from the cell-free extract of A. tumefaciens. The molecular mass of the purified enzyme estimated by HPLC was about 102 kDa, and the enzyme consisted of four subunits identical in molecular mass. The enzyme exhibited a broad absorption spectrum in the visible range with absorption maxima at 408 nm and 705 nm, and it contained cobalt and iron. The enzyme stoichiometrically catalyzed the hydration of indole-3-acetonitrile into indole-3-acetamide with a specific activity of 13.7 mol per min per mg and a Km of 7.9 microM.

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  1. Akopyan T. N., Braunstein A. E., Goryachenkova E. V. Beta-cyanoalanine synthase: purification and characterization. Proc Natl Acad Sci U S A. 1975 Apr;72(4):1617–1621. doi: 10.1073/pnas.72.4.1617. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bartel B., Fink G. R. Differential regulation of an auxin-producing nitrilase gene family in Arabidopsis thaliana. Proc Natl Acad Sci U S A. 1994 Jul 5;91(14):6649–6653. doi: 10.1073/pnas.91.14.6649. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bartling D., Seedorf M., Mithöfer A., Weiler E. W. Cloning and expression of an Arabidopsis nitrilase which can convert indole-3-acetonitrile to the plant hormone, indole-3-acetic acid. Eur J Biochem. 1992 Apr 1;205(1):417–424. doi: 10.1111/j.1432-1033.1992.tb16795.x. [DOI] [PubMed] [Google Scholar]
  4. Bartling D., Seedorf M., Schmidt R. C., Weiler E. W. Molecular characterization of two cloned nitrilases from Arabidopsis thaliana: key enzymes in biosynthesis of the plant hormone indole-3-acetic acid. Proc Natl Acad Sci U S A. 1994 Jun 21;91(13):6021–6025. doi: 10.1073/pnas.91.13.6021. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Bradford M. M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976 May 7;72:248–254. doi: 10.1016/0003-2697(76)90527-3. [DOI] [PubMed] [Google Scholar]
  6. Castric P. A., Farnden K. J., Conn E. E. Cyanide metabolism in higher plants. V. The formation of asparagine from -cyanoalanine. Arch Biochem Biophys. 1972 Sep;152(1):62–69. doi: 10.1016/0003-9861(72)90193-2. [DOI] [PubMed] [Google Scholar]
  7. Christensen A. H., Schubert K. R. Identification of a Rhizobium trifolii plasmid coding for nitrogen fixation and nodulation genes and its interaction with pJB5JI, a Rhizobium leguminosarum plasmid. J Bacteriol. 1983 Nov;156(2):592–599. doi: 10.1128/jb.156.2.592-599.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Cluness M. J., Turner P. D., Clements E., Brown D. T., O'Reilly C. Purification and properties of cyanide hydratase from Fusarium lateritium and analysis of the corresponding chy1 gene. J Gen Microbiol. 1993 Aug;139(8):1807–1815. doi: 10.1099/00221287-139-8-1807. [DOI] [PubMed] [Google Scholar]
  9. Duran R., Nishiyama M., Horinouchi S., Beppu T. Characterization of nitrile hydratase genes cloned by DNA screening from Rhodococcus erythropolis. Biosci Biotechnol Biochem. 1993 Aug;57(8):1323–1328. doi: 10.1271/bbb.57.1323. [DOI] [PubMed] [Google Scholar]
  10. Hashimoto Y., Nishiyama M., Ikehata O., Horinouchi S., Beppu T. Cloning and characterization of an amidase gene from Rhodococcus species N-774 and its expression in Escherichia coli. Biochim Biophys Acta. 1991 Feb 16;1088(2):225–233. doi: 10.1016/0167-4781(91)90058-t. [DOI] [PubMed] [Google Scholar]
  11. Ikehata O., Nishiyama M., Horinouchi S., Beppu T. Primary structure of nitrile hydratase deduced from the nucleotide sequence of a Rhodococcus species and its expression in Escherichia coli. Eur J Biochem. 1989 May 15;181(3):563–570. doi: 10.1111/j.1432-1033.1989.tb14761.x. [DOI] [PubMed] [Google Scholar]
  12. JONES E. R. H., HENBEST H. B., SMITH G. F., BENTLEY J. A. 3-indolylacetonitrile: a naturally occurring plant growth hormone. Nature. 1952 Mar 22;169(4299):485–487. doi: 10.1038/169485a0. [DOI] [PubMed] [Google Scholar]
  13. Kindl H. Oxydasen und Oxygenasen in höheren Pflanzen, I. Uber das Vorkommen von Indolyl-(3)-acetaldehydoxim und seine Bildung aus L-Tryptophan. Hoppe Seylers Z Physiol Chem. 1968 Apr;349(4):519–520. [PubMed] [Google Scholar]
  14. Klee H., Montoya A., Horodyski F., Lichtenstein C., Garfinkel D., Fuller S., Flores C., Peschon J., Nester E., Gordon M. Nucleotide sequence of the tms genes of the pTiA6NC octopine Ti plasmid: two gene products involved in plant tumorigenesis. Proc Natl Acad Sci U S A. 1984 Mar;81(6):1728–1732. doi: 10.1073/pnas.81.6.1728. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Kobayashi M., Izui H., Nagasawa T., Yamada H. Nitrilase in biosynthesis of the plant hormone indole-3-acetic acid from indole-3-acetonitrile: cloning of the Alcaligenes gene and site-directed mutagenesis of cysteine residues. Proc Natl Acad Sci U S A. 1993 Jan 1;90(1):247–251. doi: 10.1073/pnas.90.1.247. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Kobayashi M., Komeda H., Nagasawa T., Nishiyama M., Horinouchi S., Beppu T., Yamada H., Shimizu S. Amidase coupled with low-molecular-mass nitrile hydratase from Rhodococcus rhodochrous J1. Sequencing and expression of the gene and purification and characterization of the gene product. Eur J Biochem. 1993 Oct 1;217(1):327–336. doi: 10.1111/j.1432-1033.1993.tb18250.x. [DOI] [PubMed] [Google Scholar]
  17. Kobayashi M., Nagasawa T., Yamada H. Enzymatic synthesis of acrylamide: a success story not yet over. Trends Biotechnol. 1992 Nov;10(11):402–408. doi: 10.1016/0167-7799(92)90283-2. [DOI] [PubMed] [Google Scholar]
  18. MAHADEVAN S., THIMANN K. V. NITRILASE. II. SUBSTRATE SPECIFICITY AND POSSIBLE MODE OF ACTION. Arch Biochem Biophys. 1964 Jul;107:62–68. doi: 10.1016/0003-9861(64)90269-3. [DOI] [PubMed] [Google Scholar]
  19. Nagasawa T., Mauger J., Yamada H. A novel nitrilase, arylacetonitrilase, of Alcaligenes faecalis JM3. Purification and characterization. Eur J Biochem. 1990 Dec 27;194(3):765–772. doi: 10.1111/j.1432-1033.1990.tb19467.x. [DOI] [PubMed] [Google Scholar]
  20. Nagasawa T., Nanba H., Ryuno K., Takeuchi K., Yamada H. Nitrile hydratase of Pseudomonas chlororaphis B23. Purification and characterization. Eur J Biochem. 1987 Feb 2;162(3):691–698. doi: 10.1111/j.1432-1033.1987.tb10692.x. [DOI] [PubMed] [Google Scholar]
  21. Nagasawa T., Takeuchi K., Yamada H. Characterization of a new cobalt-containing nitrile hydratase purified from urea-induced cells of Rhodococcus rhodochrous J1. Eur J Biochem. 1991 Mar 28;196(3):581–589. doi: 10.1111/j.1432-1033.1991.tb15853.x. [DOI] [PubMed] [Google Scholar]
  22. Nagasawa T., Takeuchi K., Yamada H. Occurrence of a cobalt-induced and cobalt-containing nitrile hydratase in Rhodococcus rhodochrous J1. Biochem Biophys Res Commun. 1988 Sep 15;155(2):1008–1016. doi: 10.1016/s0006-291x(88)80597-7. [DOI] [PubMed] [Google Scholar]
  23. Normanly J., Cohen J. D., Fink G. R. Arabidopsis thaliana auxotrophs reveal a tryptophan-independent biosynthetic pathway for indole-3-acetic acid. Proc Natl Acad Sci U S A. 1993 Nov 1;90(21):10355–10359. doi: 10.1073/pnas.90.21.10355. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Peiser G. D., Wang T. T., Hoffman N. E., Yang S. F., Liu H. W., Walsh C. T. Formation of cyanide from carbon 1 of 1-aminocyclopropane-1-carboxylic acid during its conversion to ethylene. Proc Natl Acad Sci U S A. 1984 May;81(10):3059–3063. doi: 10.1073/pnas.81.10.3059. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Rausch V. L., Bartzack C., Catchings B., Howard B., Pruden E. Report of the topic study group on graduate education in medical technology. Am J Med Technol. 1982 Nov;48(11):935–942. [PubMed] [Google Scholar]
  26. Stevenson D. E., Feng R., Dumas F., Groleau D., Mihoc A., Storer A. C. Mechanistic and structural studies on Rhodococcus ATCC 39484 nitrilase. Biotechnol Appl Biochem. 1992 Jun;15(3):283–302. [PubMed] [Google Scholar]
  27. THIMANN K. V., MAHADEVAN S. NITRILASE. I. OCCURRENCE, PREPARATION, AND GENERAL PROPERTIES OF THE ENZYME. Arch Biochem Biophys. 1964 Apr;105:133–141. doi: 10.1016/0003-9861(64)90244-9. [DOI] [PubMed] [Google Scholar]
  28. Thimann K. V. On the Physiology of the Formation of Nodules on Legume Roots. Proc Natl Acad Sci U S A. 1936 Aug;22(8):511–514. doi: 10.1073/pnas.22.8.511. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Wright A. D., Sampson M. B., Neuffer M. G., Michalczuk L., Slovin J. P., Cohen J. D. Indole-3-Acetic Acid Biosynthesis in the Mutant Maize orange pericarp, a Tryptophan Auxotroph. Science. 1991 Nov 15;254(5034):998–1000. doi: 10.1126/science.254.5034.998. [DOI] [PubMed] [Google Scholar]
  30. Yamada T., Palm C. J., Brooks B., Kosuge T. Nucleotide sequences of the Pseudomonas savastanoi indoleacetic acid genes show homology with Agrobacterium tumefaciens T-DNA. Proc Natl Acad Sci U S A. 1985 Oct;82(19):6522–6526. doi: 10.1073/pnas.82.19.6522. [DOI] [PMC free article] [PubMed] [Google Scholar]

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