Exploitation of structural and regulatory diversity in glutamate racemases.
Glutamate racemase is an enzyme essential to the bacterial cell wall biosynthesis pathway, and has therefore been considered as a target for antibacterial drug discovery. We characterized the glutamate racemases of several pathogenic bacteria using structural and biochemical approaches. Here we describe three distinct mechanisms of regulation for the family of glutamate racemases: allosteric activation by metabolic precursors, kinetic regulation through substrate inhibition, and D-glutamate recycling using a d-amino acid transaminase. In a search for selective inhibitors, we identified a series of uncompetitive inhibitors specifically targeting Helicobacter pylori glutamate racemase that bind to a cryptic allosteric site, and used these inhibitors to probe the mechanistic and dynamic features of the enzyme. These structural, kinetic and mutational studies provide insight into the physiological regulation of these essential enzymes and provide a basis for designing narrow-spectrum antimicrobial agents.
Lundqvist
Tomas
T
AstraZeneca Global Structural Chemistry, AstraZeneca R&D Mölndal, SE-431 83, Mölndal, Sweden.
Fisher
Stewart L
SL
Kern
Gunther
G
Folmer
Rutger H A
RH
Xue
Yafeng
Y
Newton
D Trevor
DT
Keating
Thomas A
TA
Alm
Richard A
RA
de Jonge
Boudewijn L M
BL
D000494:Allosteric Regulation / Q000187:drug effects; D000495:Allosteric Site / Q000187:drug effects; D000591:Amino Acid Isomerases / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D000900:Anti-Bacterial Agents / Q000378:metabolism / Q000494:pharmacology; D001419:Bacteria / Q000201:enzymology* / Q000472:pathogenicity; D018360:Crystallography, X-Ray; D004791:Enzyme Inhibitors / Q000378:metabolism / Q000494:pharmacology; D004926:Escherichia coli / Q000201:enzymology; D016480:Helicobacter pylori / Q000201:enzymology; D007700:Kinetics; D008958:Models, Molecular; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D013329:Structure-Activity Relationship
D000900:Anti-Bacterial Agents; D004791:Enzyme Inhibitors; D011994:Recombinant Proteins; D000591:Amino Acid Isomerases; C062292:glutamate racemase