Screening for Inhibitors of Acetaldehyde Dehydrogenase (AdhE) from Enterohemorrhagic Escherichia coli (EHEC)

Abstract

Acetaldehyde dehydrogenase (AdhE) is a bifunctional acetaldehyde-coenzyme A (CoA) dehydrogenase and alcohol dehydrogenase involved in anaerobic metabolism in gram-negative bacteria. This enzyme was recently found to be a key regulator of the type three secretion (T3S) system in Escherichia coli. AdhE inhibitors can be used as tools to study bacterial virulence and a starting point for discovery of novel antibacterial agents. We developed a robust enzymatic assay, based on the acetaldehyde-CoA dehydrogenase activity of AdhE using both absorption and fluorescence detection models (Z' > 0.7). This assay was used to screen ~11,000 small molecules in 384-well format that resulted in three hits that were confirmed by resynthesis and validation. All three compounds are noncompetitive with respect to acetaldehyde and display a clear dose-response effect with hill slopes of 1-2. These new inhibitors will be used as chemical tools to study the interplay between metabolism and virulence and the role of AdhE in T3S regulation in gram-negative bacteria, and as starting points for the development of novel antibacterial agents.

Filamentation of the bacterial bi-functional alcohol/aldehyde dehydrogenase AdhE is essential for substrate channeling and enzymatic regulation

“…AdhE converts acetyl-coenzyme A to acetaldehyde and then to ethanol, in a two-step reaction that is coupled with the oxidation of two NADH molecules into NAD + . The biochemistry of AdhE enzymatic activities is well characterized 2,3 . In particular, a lot of investigations looked into this key fermentative enzyme to engineer novel ethanolproducing bacteria 4,5 or identify bacterial strains capable of growing under ethanol 6 .…”

Aldehyde-alcohol dehydrogenase undergoes structural transition to form extended spirosomes for substrate channeling

“…Previous studies also report that AdhE acts as a virulence factor in Streptococcus pneumoniae 7 and that AdhE is upregulated during infection in Salmonella typhimurium 8 . The observation that AdhE is important for virulence in several pathogens broadens its appeal as a drug target 9 . Furthermore, for industrial purpose, AdhE in fermentative bacteria is the key enzyme to produce ethanol 10 .…”

Filamentation of the bacterial bi-functional alcohol/aldehyde dehydrogenase AdhE is essential for substrate channeling and enzymatic regulation

“…AdhE converts acetyl-coenzyme A to acetaldehyde and then to ethanol, in a two-step reaction that is coupled with the oxidation of two NADH molecules into NAD + . The biochemistry of AdhE enzymatic activities is well characterized 2,3 . In particular, a lot of investigations looked into this key fermentative enzyme in order to engineer novel ethanol-producing bacteria 4,5 or identify bacterial strains capable of growing under ethanol 6 .…”