Kinetic Characterization of a Flavin-dependent Monooxygenase from the Insect Food Crop Pest, Zonocerus Variegatus

Abstract

Zonocerus variegatus, or the painted grasshopper, is a food crop pest endemic in Western and Central Africa. Agricultural industries in these regions rely heavily on natural defense mechanisms to control the grasshopper population such as plant-secreted alkaloid compounds. In recent years, the Z. variegatus population has continued to rise due to acquired resistance to alkaloids. Here we focus on the kinetic characterization of a flavin-dependent monooxygenase, ZvFMO, that catalyzes the nitrogen oxidation of many of these alkaloid compounds and confers resistance to the insect. Expression and purification of ZvFMO through a traditional E. coli expression system was successful and provided a unique opportunity to characterize the catalytic properties of an FMO from insects. ZvFMO was found to catalyze oxidation reactions of tertiary nitrogen atoms and the sulfur of cysteamine. Using stopped-flow spectroscopy, we have determined the kinetic mechanism of ZvFMO. We assessed F383 for its involvement in substrate binding, which was previously proposed, and determined that this residue does not play a major role in binding substrates. Through molecular docking, we identified N304 and demonstrated that this residue plays a role in substrate binding. The role of K215 was studied and was shown that it plays a critical role in NAD(P)H binding and cofactor selectivity.

Department(s)

Chemistry

Comments

National Science Foundation, Grant CHE 2003658

International Standard Serial Number (ISSN)

1096-0384; 0003-9861

Document Type

Article - Journal

Document Version

Citation

File Type

text

Language(s)

English

Rights

© 2024 Elsevier, All rights reserved.

Publication Date

01 Apr 2024

PubMed ID

38430968

Link to Full Text

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