Essentiality and functional analysis of type I and type III pantothenate kinases of Mycobacterium tuberculosis

Abstract

Pantothenate kinase, an essential enzyme in bacteria and eukaryotes, is involved in catalysing the first step of conversion of pantothenate to coenzyme A (CoA). Three isoforms (type I, II and III) of this enzyme have been reported from various organisms, which can be differentiated from each other on the basis of their biochemical and structural characteristics. Though most bacteria carry only one of the isoforms of pantothenate kinases, some of them possess two isoforms. The physiological relevance of the presence of two types of isozymes in a single organism is not clear.Mycobacterium tuberculosis,an intracellular pathogen, possesses two isoforms of pantothenate kinases (CoaA and CoaX) belonging to type I and III. In order to determine which pantothenate kinase is essential in mycobacteria, we performed gene inactivation ofcoaAandcoaXofM. tuberculosisindividually. It was found that coaAcould only be inactivated in the presence of an extra copy of the gene, whilecoaXcould be inactivated in the wild-type cells, proving that CoaA is the essential pantothenate kinase inM. tuberculosis. Additionally, thecoaAgene ofM. tuberculosiswas able to complement a temperature-sensitivecoaAmutant ofEscherichia coliat a non-permissive temperature whilecoaXcould not. ThecoaXdeletion mutant showed no growth defectsin vitro, in macrophages or in mice. Taken together, our data suggest that CoaX, which is essential inBacillus anthracisand thus had been suggested to be a drug target in this organism, might not be a valid target inM. tuberculosis. We have established that the type I isoform, CoaA, is the essential pantothenate kinase inM. tuberculosisand thus can be explored as a drug target.