A novel inhibitor of Mycobacterium tuberculosis pantothenate synthetase.
Pantothenate synthetase (PS; EC 6.3.2.1), encoded by the panC gene, catalyzes the essential adenosine triphosphate (ATP)-dependent condensation of D-pantoate and beta-alanine to form pantothenate in bacteria, yeast, and plants; pantothenate is a key precursor for the biosynthesis of coenzyme A (CoA) and acyl carrier protein (ACP). Because the enzyme is absent in mammals and both CoA and ACP are essential cofactors for bacterial growth, PS is an attractive chemotherapeutic target. An automated high-throughput screen was developed to identify drugs that inhibit Mycobacterium tuberculosis PS. The activity of PS was measured spectrophotometrically through an enzymatic cascade involving myokinase, pyruvate kinase, and lactate dehydrogenase. The rate of PS ATP utilization was quantitated by the reduction of absorbance due to the oxidation of NADH to NAD+ by lactate dehydrogenase, which allowed for an internal control to detect interference from compounds that absorb at 340 nm. This coupled enzymatic reaction was used to screen 4080 compounds in a 96-well format. This discussion describes a novel inhibitor of PS that exhibits potential as an antimicrobial agent.
White
E Lucile
EL
Southern Research Institute, Birmingham, AL 35205-5305, USA. white@sri.org
Southworth
Kristen
K
Ross
Larry
L
Cooley
Sara
S
Gill
Rachel B
RB
Sosa
Melinda Ingrum
MI
Manouvakhova
Anna
A
Rasmussen
Lynn
L
Goulding
Celia
C
Eisenberg
David
D
Fletcher
Thomas M
TM
D004791:Enzyme Inhibitors / Q000032:analysis / Q000737:chemistry / Q000494:pharmacology*; D008958:Models, Molecular; D009169:Mycobacterium tuberculosis / Q000187:drug effects / Q000201:enzymology*; D009257:Nafronyl / Q000737:chemistry; D010453:Peptide Synthases / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000378:metabolism; D015203:Reproducibility of Results
D004791:Enzyme Inhibitors; D009257:Nafronyl; D010453:Peptide Synthases; C022318:pantothenate synthetase