Biochemical Characterization of a Putative Glycerol-3- Phosphate Acyltransferase From Mycobacterium tuberculosis

Jamison Law, Purdue University


Latent tuberculosis affects over two billion people and is caused by dormant Mycobacterium tuberculosis (Mtb) that is phenotypically tolerant to antibiotics and relies upon triacylglycerol (TAG) accumulation for survival in the host cell. The mycobacterial glycerol-3-phosphate acyltransferase (mGPAT) is responsible for catalyzing the first step of the TAG biosynthetic pathway, but it has not been studied. Therefore, we cloned the open reading frame of a putative mGPAT and expressed it in Escherichia coli to study its function. We observed that cell lysates expressing mGPAT displayed increased radiolabel accumulation in phospholipids and pure mGPAT formed an unknown product. These results suggest that mGPAT increased the formation of precursor substrates that were further utilized in lipid biosynthesis. Significant differences were observed in the metabolic incorporation of 14C-palmitate and 14C-acetate into total lipids and lipid subclasses when E. coli was expressing mGPAT. This suggests that mGPAT affects E. coli’s preference for endogenous versus exogenous sources of fatty acid for incorporation into phospholipids. When cultured in media supplemented with fatty acid, E. coli expressing mGPAT exhibited significantly higher growth compared to E. coli lacking mGPAT during exponential and stationary phase. E. coli expressing mGPAT cultured in nutrient-rich media displayed lower growth during exponential phase and higher growth during stationary phase as compared to E. coli lacking mGPAT. These results suggest that mGPAT enhances the ability of E. coli to utilize exogenously provided long-chain fatty acids for membrane lipid biosynthesis under nutrient-limiting conditions, which is similar to the conditions that Mtb experiences inside the human body.




Daniel, Purdue University.

Subject Area

Molecular biology|Microbiology|Biochemistry

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