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VIEW ARTICLE   |    DOI: 10.1094/MPMI-1-128

Properties of Two Enzymes Involved in the Biosynthesis of the Fungal Pathogenicity Factor HC-Toxin. Jonathan D. Walton. Department of Energy-Plant Research Laboratory, Michigan State University, E. Lansing, MI 48824 U.S.A.. Frank R. Holden. Department of Energy-Plant Research Laboratory, Michigan State University, E. Lansing, MI 48824 U.S.A.. MPMI 1:128-134. Accepted 22 February 1988. Copyright 1988 The American Phytopathological Society.

Host-selective toxins are chemical mediators of the plant-pathogen interactions in which they occur. In the fungal maize pathogen Cochliobolus carbonum race 1, a single genetic locus controls production of its host-selective toxin, HC-toxin. This genetic locus is associated with two separable enzymatic activities from C. carbonum, which have the properties expected of enzymes involved in the biosynthesis of HC-toxin (Walton, Proc. Natl. Acad. Sci. USA 84:8444, 1987). These enzymes have been purified and characterized. One enzyme, called HTS-1, activates l-proline by adenosine triphosphate/inorganic pyrophosphate exchange, binds l-proline as the thioester, and epimerizes l-proline to d-proline, the isomer of proline found in HC-toxin. The second enzyme, called HTS-2, activates both l-alanine and d-alanine, forms the thioesters of each amino acid, and epimerizes l-alanine to d-alanine, but not d-alanine to l-alanine. The two enzymes were purified by precipitation with ammonium sulfate and anion exchange, gel filtration, and hydroxyapatite chromatography. By HPLC gel filtration HTS-1 and HTS-2 both have apparent molecular weights of 310,000, but by sodium dodecyl sulfate-PAGE the apparent molecular weights are 220,000 and 160,000, respectively. HTS-1 has a Michaelis constant for l-proline of 17.6 mM. HTS-2 has Michaelis constants of 3.4 and 101 mM for d-alanine and l-alanine, respectively, but the maximum velocity for l-alanine is three times that for d-alanine and approximately equal to that of HTS-1 for l-proline. The results in this paper demonstrate the involvement of HTS-1 and HTS-2 in the biosynthesis of HC-toxin and describe methods to purify the enzymes, opening an avenue to elucidate one aspect of the molecular biology of the interaction between C. carbonum race 1 and maize.

Additional Keywords: cyclic peptide biosynthesis, maize.