Dafeng Liu, Cai Yuan, Chenyun Guo, Mingdong Huang, Donghai Lin
Protein expression and purification 2024 FebMycobacterium tuberculosis (Mtb) is a crucial and highly destructive intracellular pathogen responsible for causing tuberculosis (TB). The emergence and dissemination of multi-drug resistant Mtb has further aggravated the TB crisis, leading to high mortality. Mtb FadD2 is a fatty acyl-coenzyme A (CoA) synthetase that modifies the cell envelope and plays an important role in reducing Mtb susceptibility to pyrazinoic acid (POA). However, the functional mechanism of Mtb FadD2 remains poorly understood. Here, we successfully expressed, purified and obtained monomeric FadD2 by using buffer (500 mM NaCl, 20 mM Tris-HCl, pH7.4 and 5 % glycerol). Palmitate was found to be the optimal substrate for FadD2. Fatty acyl-CoA synthetase activity reached maximum at 450 μM palmitate, and the Km value was 318.2 μM for palmitate. The results of mutation experiments indicated the critical role of T370 and K551 in the enzymatic activity of FadD2. Our work provides a guideline and concept for the development of novel drugs against Mtb. Copyright © 2023 Elsevier Inc. All rights reserved.
Dafeng Liu, Cai Yuan, Chenyun Guo, Mingdong Huang, Donghai Lin. Recombinant expression and functional characterization of FadD2 protein in Mycobacterium tuberculosis. Protein expression and purification. 2024 Feb;214:106377
PMID: 37813293
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