In vitro kinetic analysis of substrate specificity in enterobactin biosynthetic lower pathway enzymes provides insight into the biochemical function of the hot dog-fold thioesterase EntH.
The Escherichia coli siderophore enterobactin is assembled from 2,3-dihydroxybenzoate (2,3-DHB) and l-serine by the nonribosomal peptide synthetases EntB and EntF. The processive thiol-template strategy used can be sabotaged by EntB misacylation. Through in vitro kinetic analysis, we demonstrate two potential routes to EntB misacylation and provide evidence for two mechanisms by which the hot dog-fold thioesterase EntH can potentially prevent or reverse EntB misacylation.
Chen
Danqi
D
Department of Chemistry and Chemical Biology, University of New Mexico, Albuquerque, New Mexico 87131, USA.
Wu
Rui
R
Bryan
Tyrel L
TL
Dunaway-Mariano
Debra
D
D000214:Acyl Coenzyme A / Q000378:metabolism; D001577:Benzophenones / Q000378:metabolism; D002384:Catalysis; D004758:Enterobactin / Q000096:biosynthesis* / Q000737:chemistry; D004926:Escherichia coli / Q000201:enzymology*; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000378:metabolism*; D020035:Holoenzymes / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D007700:Kinetics; D017433:Protein Structure, Secondary; D021241:Spectrometry, Mass, Electrospray Ionization; D013379:Substrate Specificity; D013696:Temperature; D013869:Thiolester Hydrolases / Q000737:chemistry* / Q000378:metabolism*; D013997:Time Factors
D000214:Acyl Coenzyme A; D001577:Benzophenones; D029968:Escherichia coli Proteins; D020035:Holoenzymes; D004758:Enterobactin; C527558:EntH protein, E coli; D013869:Thiolester Hydrolases; C121479:2,4-dihydroxybenzophenone