Role for malonyl coenzyme A:acyl carrier protein transacylase (MCAT) in the growth-inhibitory effect of the calmodulin antagonist trifluoperazine in Mycobacterium bovis BCG

Objectives: To determine whether the fatty acid synthesis enzyme malonyl coenzyme A:acyl carrier protein transacylase (MCAT) is involved in the growth-inhibitory effect of trifluoperazine in the tubercle bacillus Mycobacterium bovis BCG. Methods: BCG was grown in liquid culture with various concentrations of trifluoperazine and growth was monitored by OD measurement. To determine the effect of trifluoperazine on MCAT protein level, total protein was extracted from BCG cultures and was analysed by 2D gel electrophoresis and western blot. To confirm trifluoperazine-dependent reduction in the MCAT protein level, two BCG strains overexpressing MCAT at a low and high constitutive level were similarly tested. The synergic effect of trifluoperazine and isoniazid was tested at sub-MIC levels in liquid cultures. Results: Trifluoperazine inhibition of growth correlates with reduction in the steady-state level of MCAT protein. Overexpression of MCAT confers resistance to trifluoperazine. Trifluoperazine acts synergically (albeit weakly) with isoniazid and no resistance towards isoniazid alone was observed due to overexpression of MCAT. This suggests MCAT to be a specific target of trifluoperazine. Conclusion: These results indicate MCAT as a target of trifluoperazine and provide an explanation for the inhibitory effect of trifluoperazine on mycobacterial lipid synthesis observed earlier. This makes MCAT a potential target for new antimycobacterials.