Poster: Hormones

Abs # 585: Analysis of the tryptophan-dependent indole-3-acetic acid biosynthesis pathway in maize endosperm

Presenter:	Hendrickson, Angela M, hend0409@tc.umn.edu
Authors	Hendrickson, Angela M (A)   Cohen, Jerry D (A)
Affiliations:	(A): University of Minnesota

Zea mays endosperm produces very high levels of endogenous indole-3-acetic acid (IAA), which is an important hormone in developing maize embryos and is stored as conjugates in the endosperm where it is later used during the early stages of seedling growth. Although tryptophan-dependent IAA biosynthesis pathways have been widely studied and confirmed by several groups to be responsible for the major fraction of IAA production in maturing maize endosperm tissue, a complete pathway has yet to be isolated from any higher plant. This work focuses on identifying intermediates and isolating enzymes in the tryptophan-dependent pathway using maize endosperm. Each purification step for the identification of intermediate enzymes is conducted by feeding [²H₅]tryptophan to enzyme preparation from maize endosperm, and then quantifying the amount of [²H₅]IAA symthesized using gas chromatography-mass spectometry (GC-MS) with [¹³C₆]IAA as an internal standard. The complete set of enzymes appear to be membrane associated. The membrane-bound enzymes are thus solubolized using DDM and TritonX-100, further purified, and then assayed for conversion activity. Intermediate compounds were partially characterized by "cold trapping" methods and we are currently developing supplemental methods using radioHPLC and LC-MS procedures. This work was supported by grand DE-A102-94ER20153 from the U.S. Department of Energy, and by a USDA National Needs fellowship to AMH.