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J. Biol. Chem., Vol. 283, Issue 3, 1660-1669, January 18, 2008

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Structural and Functional Characterization of the TRI101 Trichothecene 3-O-Acetyltransferase from Fusarium sporotrichioides and Fusarium graminearum

KINETIC INSIGHTS TO COMBATING FUSARIUM HEAD BLIGHT^*

Graeme S. Garvey, Susan P. McCormick, and Ivan Rayment¹

From the Department of Biochemistry, University of Wisconsin, Madison, Wisconsin 53706 and the Mycotoxin Research Unit, USDA/ARS, National Center for Agricultural Utilization Research, Peoria, Illinois 61604

Fusarium head blight (FHB) is a plant disease with serious economic and health impacts. It is caused by fungal species belonging to the genus Fusarium and the mycotoxins they produce. Although it has proved difficult to combat this disease, one strategy that has been examined is the introduction of an indigenous fungal protective gene into cereals such as wheat barley and rice. Thus far the gene of choice has been tri101 whose gene product catalyzes the transfer of an acetyl group from acetyl coenzyme A to the C3 hydroxyl moiety of several trichothecene mycotoxins. In vitro this has been shown to reduce the toxicity of the toxins by 100-fold but has demonstrated limited resistance to FHB in transgenic cereal. To understand the molecular basis for the differences between in vitro and in vivo resistance the three-dimensional structures and kinetic properties of two TRI101 orthologs isolated from Fusarium sporotrichioides and Fusarium graminearum have been determined. The kinetic results reveal important differences in activity of these enzymes toward B-type trichothecenes such as deoxynivalenol. These differences in activity can be explained in part by the three-dimensional structures for the ternary complexes for both of these enzymes with coenzyme A and trichothecene mycotoxins. The structural and kinetic results together emphasize that the choice of an enzymatic resistance gene in transgenic crop protection strategies must take into account the kinetic profile of the selected protein.

Received for publication, July 13, 2007 , and in revised form, October 1, 2007.

The atomic coordinates and structure factors (code 2ZBA, 2RKT, 3B30, 2RKV, and 3B2S) have been deposited in the Protein Data Bank, Research Collaboratory for Structural Bioinformatics, Rutgers University, New Brunswick, NJ (http://www.rcsb.org/).

^* This work was supported by National Institutes of Health Grant AR35186 (to I. R.) and the United States Department of Agriculture under Agreement FY06-RA-098. This is a cooperative project with the United States Wheat & Barley Scab Initiative. Use of the Structural Biology BM19 beamline Argonne National Laboratory Advanced Photon Source was supported by the United States Department of Energy, Office of Energy Research Contract W-31-109-ENG-38. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

The on-line version of this article (available at http://www.jbc.org) contains supplemental Figs. S1-S3.

¹ To whom correspondence should be addressed: Dept. of Biochemistry, 433 Babcock Dr., Madison, WI 53706. Tel.: 608-262-0437; Fax: 608-262-1319; E-mail: ivan_rayment{at}biochem.wisc.edu.

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