Systems and strategies for plant protein expression.

Autor:	Zmuda AJ; Department of Plant and Microbial Biology, University of Minnesota, St. Paul, MN, United States., Niehaus TD; Department of Plant and Microbial Biology, University of Minnesota, St. Paul, MN, United States. Electronic address: tniehaus@umn.edu.
Jazyk:	angličtina
Zdroj:	Methods in enzymology [Methods Enzymol] 2023; Vol. 680, pp. 3-34. Date of Electronic Publication: 2022 Sep 02.
DOI:	10.1016/bs.mie.2022.08.002
Abstrakt:	At least a quarter of the protein-encoding genes in plant genomes are predicted to encode enzymes for which no physiological function is known. Determining functions for these uncharacterized enzymes is key to understanding plant metabolism. Functional characterization typically requires expression and purification of recombinant enzymes to be used in enzyme assays and/or for protein structure elucidation studies. Here, we describe several practical considerations used to improve the heterologous expression and purification of Arabidopsis thaliana and Zea mays NAD(P)HX dehydratase (NAXD) and NAD(P)HX epimerase (NAXE), two enzymes that are involved in repair of chemically damaged NAD(P)H cofactors. We provide protocols for transit peptide prediction and construct design, expression in Escherichia coli, and purification of NAXD and NAXE. Many of these strategies are generally applicable to the purification of any plant protein. (Copyright © 2023 Elsevier Inc. All rights reserved.)
Databáze:	MEDLINE
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