Analysis of coenzyme A activated compounds in actinomycetes.

CABRUJA M, , LYONNET BB, MILLÁN G, GRAMAJO H, GAGO G

APPLIED MICROBIOLOGY AND BIOTECHNOLOGY

SPRINGER

Lugar: Berlin; Año: 2016

0175-7598

Abstract Acyl-CoAs are crucial compounds involved in essentialmetabolic pathways such as the Krebs cycle and lipid,carbohydrate, and amino acid metabolisms, and they are alsokey signalmolecules involved in the transcriptional regulationof lipid biosynthesis inmany organisms. In this study, we tookadvantage of the high selectivity of mass spectrometry anddeveloped an ion-pairing reverse-phase high-pressure liquidchromatography electrospray ionization high-resolution massspectrometry (IP-RP-HPLC/ESI-HRMS)method to carry on acomprehensive analytical determination of the wide range offatty acyl-CoAs present in actinomycetes. The advantage ofusing a QTOF spectrometer resides in the excellent mass accuracyover a wide dynamic range and measurements of thetrue isotope pattern that can be used for molecular formulaelucidation of unknown analytes. As a proof of concept, weused this assay to determine the composition of the fatty acyl-CoA pools in Mycobacterium, Streptomyces, andCorynebacterium species, revealing an extraordinary differencein fatty acyl-CoA amounts and species distribution betweenthe three genera and between the two species ofmycobacteria analyzed, including the presence of differentchain-length carboxy-acyl-CoAs, key substrates of mycolicacid biosynthesis. The method was also used to analyze theimpact of two fatty acid synthase inhibitors on the acyl-CoAprofile of Mycobacterium smegmatis, which showed someunexpected low levels of C24 acyl-CoAs in the isoniazidtreatedcells. This robust, sensitive, and reliable methodshould be broadly applicable in the studies of the wide rangeof bacteria metabolisms in which acyl-CoA moleculesparticipate.