A method for identification of inhibitors of the phosphorylation reactions of bacterial response regulator proteins using (31)P nuclear magnetic resonance spectroscopy.
Bacterial response regulators are attractive targets for antibacterial drug development, yet random screening against these targets has failed as yet to identify chemicals that constitute viable leads. Alternative methods to provide leads for drug development based on identification and optimization of low affinity ligands from NMR screens have been described. However, leads from these processes still require verification in a bioassay, which is often problematic if compounds have unfavorable optical and solubility properties. A simple method, based on using NMR to observe the activity of the target, is described. It has the advantages of being able to characterize both low affinity leads and a wider selection of compounds in a structure activity relationships series, without the problems affecting a fluorescence assay. In this example we use (31)P to monitor the turnover of a bacterial response regulator, but the generic approach could be applied to other nuclei and thus a range of biological systems.
Hubbard
J A
JA
Computational and Structural Sciences, GlaxoSmithKline, Medicines Research Centre, Gunnels Wood Road, Stevenage, Hertfordshire SG1 2NY, United Kingdom.
MacLachlan
L K
LK
Johnson
P
P
Findlow
S C
SC
Ladds
J C
JC
Lewis
C J
CJ
Carter
P S
PS
Jones
J J
JJ
Fosberry
A P
AP
D000900:Anti-Bacterial Agents / Q000378:metabolism / Q000494:pharmacology; D001426:Bacterial Proteins / Q000187:drug effects / Q000302:isolation & purification / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism*; D002384:Catalysis; D004492:Edetic Acid / Q000378:metabolism; D008024:Ligands; D008274:Magnesium / Q000378:metabolism*; D008565:Membrane Proteins / Q000302:isolation & purification / Q000378:metabolism*; D000072236:Methyl-Accepting Chemotaxis Proteins; D008970:Molecular Weight; D019906:Nuclear Magnetic Resonance, Biomolecular / Q000379:methods*; D010759:Phosphorus Isotopes / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects
D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D008024:Ligands; D008565:Membrane Proteins; D000072236:Methyl-Accepting Chemotaxis Proteins; D010759:Phosphorus Isotopes; D004492:Edetic Acid; D008274:Magnesium; D002118:Calcium