skip to main content
OSTI.GOV title logo U.S. Department of Energy
Office of Scientific and Technical Information

Title: An in vitro system from maize seedlings for tryptophan-independent indole-3-acetic acid biosynthesis

Abstract

The enzymatic synthesis of indole-3-acetic acid (IAA) from indole by an in vitro preparation from maize (Zea mays L.) that does not use tryptophan (Trp) as an intermediate is described. Light-grown seedlings of normal maize and the maize mutant orange pericarp were shown to contain the necessary enzymes to convert [{sup 14}C]indole to IAA. The reaction was not inhibited by unlabeled Trp and neither [{sup 14}C]Trp nor [{sup 14}C]serine substituted for [{sup 14}C]indole in this in vitro system. The reaction had a pH optimum greater than 8.0, required a reducing environment, and had an oxidation potential near that of ascorbate. The results obtained with this in vitro enzyme preparation provide strong, additional evidence for the presence of a Trp-independent IAA biosynthesis pathway in plants.

Authors:
 [1];  [2];  [3]
  1. Swedish Univ. of Agricultural Sciences, Umeaa (Sweden). Dept. of Forest Genetics and Plant Physiology
  2. Univ. of Maryland, College Park, MD (United States). Dept. of Plant Biology
  3. Dept. of Agriculture, Beltsville, MD (United States)
Publication Date:
Sponsoring Org.:
USDOE, Washington, DC (United States)
OSTI Identifier:
305473
Resource Type:
Journal Article
Resource Relation:
Journal Name: Plant Physiology (Bethesda); Journal Volume: 119; Journal Issue: 1; Other Information: PBD: Jan 1999
Country of Publication:
United States
Language:
English
Subject:
55 BIOLOGY AND MEDICINE, BASIC STUDIES; MAIZE; ACETIC ACID; BIOSYNTHESIS; SEEDLINGS; INDOLES; BIOLOGICAL PATHWAYS; ENZYME ACTIVITY

Citation Formats

Oestin, A., Ilic, N., and Cohen, J.D. An in vitro system from maize seedlings for tryptophan-independent indole-3-acetic acid biosynthesis. United States: N. p., 1999. Web. doi:10.1104/pp.119.1.173.
Oestin, A., Ilic, N., & Cohen, J.D. An in vitro system from maize seedlings for tryptophan-independent indole-3-acetic acid biosynthesis. United States. doi:10.1104/pp.119.1.173.
Oestin, A., Ilic, N., and Cohen, J.D. 1999. "An in vitro system from maize seedlings for tryptophan-independent indole-3-acetic acid biosynthesis". United States. doi:10.1104/pp.119.1.173.
@article{osti_305473,
title = {An in vitro system from maize seedlings for tryptophan-independent indole-3-acetic acid biosynthesis},
author = {Oestin, A. and Ilic, N. and Cohen, J.D.},
abstractNote = {The enzymatic synthesis of indole-3-acetic acid (IAA) from indole by an in vitro preparation from maize (Zea mays L.) that does not use tryptophan (Trp) as an intermediate is described. Light-grown seedlings of normal maize and the maize mutant orange pericarp were shown to contain the necessary enzymes to convert [{sup 14}C]indole to IAA. The reaction was not inhibited by unlabeled Trp and neither [{sup 14}C]Trp nor [{sup 14}C]serine substituted for [{sup 14}C]indole in this in vitro system. The reaction had a pH optimum greater than 8.0, required a reducing environment, and had an oxidation potential near that of ascorbate. The results obtained with this in vitro enzyme preparation provide strong, additional evidence for the presence of a Trp-independent IAA biosynthesis pathway in plants.},
doi = {10.1104/pp.119.1.173},
journal = {Plant Physiology (Bethesda)},
number = 1,
volume = 119,
place = {United States},
year = 1999,
month = 1
}
  • We are attempting to determine when seedlings of Zea mays sweet corn, var. Silver Queen begin de novo biosynthesis of tryptophan and indole-3-acetic acid (IAA). We wish to use the general precursor, deuterium labeled water, to minimize assumptions as to the biosynthetic route. Protium in positions 2, 4, 5, 6 7 of the indole ring are non-exchangeable. IAA and tryptophan synthesized via the shikimic acid pathway would contain deuterium in one or more of these positions . The protium on the indene nitrogen, the carboxyl, the amino group, or the protium alpha to the carboxyl exchange readily and so aremore » removed prior to analysis by base catalyzed exchange. The IAA, or trypotophan, is then purified by DEAE, Dowex 50, and two HPLC steps. IAA is methylated with diazomethane and analyzed by GC/MS. Trytophan is acetylated with triethylamine-acetic anhydride and then methylated with diazomethane and analyzed by GC/MS. Results of these studies on plants grown for varying lengths of time and under various light and nutrient conditions will be reported.« less
  • The authors present evidence that the role of tryptophan and other potential intermediates in the pathways that could lead to indole derivatives needs to be reexamined. Two lines of Lemna gibba were tested for uptake of ({sup 15}N-indole)-labeled tryptophan isomers and incorporation of that label into free indole-3-acetic acid (IAA). Both lines required levels of L-({sup 15}N)tryptophan 2 to 3 orders of magnitude over endogenous levels in order to obtain measurable incorporation of label into IAA. Labeled L-tryptophan was extractable from plant tissue after feeding and showed no measurable isomerization into D-tryptophan. D-({sup 15}N)trytophan supplied to Lemna at rates ofmore » approximately 400 times excess of endogenous D-tryptophan levels (to yield an isotopic enrichment equal to that which allowed detection of the incorporation of L-tryptophan into IAA), did not result in measurable incorporation of label into free IAA. These results demonstrate that L-tryptophan is a more direct precursor to IAA than the D isomer and suggest (a) that the availability of tryptophan in vivo is not a limiting factor in the biosynthesis of IAA, thus implying that other regulatory mechanisms are in operation and (b) that L-tryptophan also may not be a primary precursor to IAA in plants.« less
  • Either 5-(/sup 3/H)indole-3-acetic acid (IAA) or 5-(/sup 3/H)indole-3-acetyl-myoinositol was applied to the endosperm of kernels of dark-grown Zea mays seedlings. The distribution of total radioactivity, radiolabeled indole-3-acetic acid, and radiolabeled ester conjugated indole-3-acetic acid, in the shoots was then determined. Differences were found in the distribution and chemical form of the radiolabeled indole-3-acetic acid in the shoot depending upon whether 5-(/sup 3/H)indole-3-acetic acid or 5-(/sup 3/H)indole-3-acetyl-myo-inositol was applied to the endosperm. We demonstrated that indole-3-acetyle-myo-inositol applied to the endosperm provides both free and ester conjugated indole-3-acetic acid to the mesocotyl and coleoptile. Free indole-3-acetic acid applied to the endosperm suppliesmore » some of the indole-3-acetic acid in the mesocotyl but essentially no indole-3-acetic acid to the coleoptile or primary leaves. It is concluded that free IAA from the endosperm is not a source of IAA for the coleoptile. Neither radioactive indole-3-acetyl-myo-inositol nor IAA accumulates in the tip of the coleoptile or the mesocotyl node and thus these studies do not explain how the coleoptile tip controls the amount of IAA in the shoot.« less