Crystallographic structure of ChitA, a glycoside hydrolase family 19, plant class IV chitinase from Zea mays

Authors

  • Marcia M. Chaudet,

    1. Department of Biology, University of Waterloo, Waterloo, Ontario, Canada
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  • Todd A. Naumann,

    1. US Department of Agriculture–Agricultural Research Service–National Center for Agricultural Utilization Research (USDA–ARS–NCAUR), Bacterial Foodborne Pathogens and Mycology Research Unit, Peoria, Illnois
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  • Neil P.J. Price,

    1. US Department of Agriculture–Agricultural Research Service–National Center for Agricultural Utilization Research (USDA–ARS–NCAUR), Renewable Product Technology Research Unit, Peoria, Illinois
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  • David R. Rose

    Corresponding author
    1. Department of Biology, University of Waterloo, Waterloo, Ontario, Canada
    • Correspondence to: David R. Rose, Department of Biology, University of Waterloo, 200 University Avenue West, Waterloo, Ontario, Canada N2L 3G1. E-mail: drrose@uwaterloo.ca

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  • Mention of any trade names or commercial products in this publication is solely for the purpose of providing specific information and does not imply recommendation or endorsement by the U.S. Department of Agriculture.

ABSTRACT

Maize ChitA chitinase is composed of a small, hevein-like domain attached to a carboxy-terminal chitinase domain. During fungal ear rot, the hevein-like domain is cleaved by secreted fungal proteases to produce truncated forms of ChitA. Here, we report a structural and biochemical characterization of truncated ChitA (ChitA ΔN), which lacks the hevein-like domain. ChitA ΔN and a mutant form (ChitA ΔN-EQ) were expressed and purified; enzyme assays showed that ChitA ΔN activity was comparable to the full-length enzyme. Mutation of Glu62 to Gln (ChitA ΔN-EQ) abolished chitinase activity without disrupting substrate binding, demonstrating that Glu62 is directly involved in catalysis. A crystal structure of ChitA ΔN-EQ provided strong support for key roles for Glu62, Arg177, and Glu165 in hydrolysis, and for Ser103 and Tyr106 in substrate binding. These findings demonstrate that the hevein-like domain is not needed for enzyme activity. Moreover, comparison of the crystal structure of this plant class IV chitinase with structures from larger class I and II enzymes suggest that class IV chitinases have evolved to accommodate shorter substrates.

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