A fragment-based approach to understanding inhibition of 1-deoxy-D-xylulose-5-phosphate reductoisomerase.
The inhibition of 1-deoxy-D-xylulose-5-phosphate reductoisomerase (DXR) by fosmidomycin was studied by using a kinetic assay based on the consumption of NADPH and synthetic substrate. Fosmidomycin is a slow tight-binding inhibitor of DXR that shows strong negative cooperativity (absolute value(h) = 0.3) in binding. Cooperativity is displayed during the initial (weak, K0.5 = 10 microM) binding event and does not change as the binding tightens to the equilibrium value of 0.9 nM over a period of seconds to minutes. A series of fosmidomycin fragments was examined, but all showed much weaker inhibition, in the mM range. A series of cyclic fosmidomycin analogues was also synthesised and tested, but these showed high-microM binding at best. None of the synthetic compounds showed time-dependent inhibition. We concluded that the slow tight-binding behaviour, and perhaps also cooperativity, are mediated by significant reorganisation of the active site upon fosmidomycin binding. This makes the rational design of new inhibitors of DXR difficult at best.
Mercklé
Ludovic
L
School of Chemistry, University of Bristol, Cantock's Close, Clifton, Bristol, BS8 1TS, UK.
de Andrés-Gómez
Ana
A
Dick
Bethany
B
Cox
Russell J
RJ
Godfrey
Christopher R A
CR
D019747:Aldose-Ketose Isomerases / Q000037:antagonists & inhibitors* / Q000737:chemistry* / Q000378:metabolism; D000900:Anti-Bacterial Agents / Q000737:chemistry / Q000494:pharmacology*; D001665:Binding Sites; D004791:Enzyme Inhibitors / Q000737:chemistry / Q000494:pharmacology*; D004926:Escherichia coli / Q000201:enzymology; D005578:Fosfomycin / Q000031:analogs & derivatives* / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology; D007700:Kinetics; D008958:Models, Molecular; D009097:Multienzyme Complexes / Q000037:antagonists & inhibitors* / Q000737:chemistry* / Q000378:metabolism; D010088:Oxidoreductases / Q000037:antagonists & inhibitors* / Q000737:chemistry* / Q000378:metabolism
D000900:Anti-Bacterial Agents; D004791:Enzyme Inhibitors; D009097:Multienzyme Complexes; D005578:Fosfomycin; C024640:fosmidomycin; D010088:Oxidoreductases; C114231:1-deoxy-D-xylulose 5-phosphate reductoisomerase; D019747:Aldose-Ketose Isomerases