title: Mycobacterium tuberculosis appears to lack alpha-ketoglutarate dehydrogenase and encodes pyruvate dehydrogenase in widely separated genes.
abstract: Mycobacterium tuberculosis (Mtb) persists for prolonged periods in macrophages, where it must adapt to metabolic limitations and oxidative/nitrosative stress. However, little is known about Mtb's intermediary metabolism or antioxidant defences. We recently identified a peroxynitrite reductase-peroxidase complex in Mtb that included products of the genes sucB and lpd, which are annotated to encode the dihydrolipoamide succinyltransferase (E2) and lipoamide dehydrogenase (E3) components of alpha-ketoglutarate dehydrogenase (KDH). However, we could detect no KDH activity in Mtb lysates, nor could we reconstitute KDH by combining the recombinant proteins SucA (annotated as the E1 component of KDH), SucB and Lpd. We therefore renamed the sucB product dihydrolipoamide acyltransferase (DlaT). Mtb lysates contained pyruvate dehydrogenase (PDH) activity, which was lost when the dlaT gene (formerly, sucB) was disrupted. Purification of PDH from Mtb yielded AceE, annotated as an E1 component of PDH, along with DlaT and Lpd. Moreover, anti-DlaT antibody coimmunoprecipitated AceE. Finally, recombinant AceE, DlaT and Lpd, although encoded by genes that are widely separated on the chromosome, reconstituted PDH in vitro with Km values typical of bacterial PDH complexes. In sum, Mtb appears to lack KDH. Instead, DlaT and Lpd join with AceE to constitute PDH.
authors.affiliation: Department of Microbiology and Immunology, Weill Medical College of Cornell University, 1300 York Avenue, New York, NY 10021, USA.
authors.forename: Jing
authors.initials: J
authors.lastname: Tian
authors.forename: Ruslana
authors.initials: R
authors.lastname: Bryk
authors.forename: Shuangping
authors.initials: S
authors.lastname: Shi
authors.forename: Hediye
authors.initials: H
authors.lastname: Erdjument-Bromage
authors.forename: Paul
authors.initials: P
authors.lastname: Tempst
authors.forename: Carl
authors.initials: C
authors.lastname: Nathan
mesh_terms: D000217:Acyltransferases / Q000235:genetics / Q000378:metabolism; D008058:Dihydrolipoamide Dehydrogenase / Q000235:genetics / Q000378:metabolism; D007655:Ketoglutarate Dehydrogenase Complex / Q000378:metabolism*; D009169:Mycobacterium tuberculosis / Q000201:enzymology* / Q000235:genetics; D011768:Pyruvate Dehydrogenase Complex / Q000235:genetics* / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism
chemical_list: D011768:Pyruvate Dehydrogenase Complex; D011994:Recombinant Proteins; D007655:Ketoglutarate Dehydrogenase Complex; D008058:Dihydrolipoamide Dehydrogenase; D000217:Acyltransferases; C033588:dihydrolipoamide acyltransferase