[Show abstract][Hide abstract] ABSTRACT: Acyl carrier proteins play a central role in metabolism by transporting substrates in a wide variety of pathways including the biosynthesis of fatty acids and polyketides. However, despite their importance, there is a paucity of direct structural information concerning the interaction of ACPs with enzymes in these pathways. Here we report the structure of an acyl-ACP substrate bound to the Escherichia coli fatty acid biosynthesis enoyl reductase enzyme (FabI), based on a combination of x-ray crystallography and molecular dynamics simulation. The structural data are in agreement with kinetic studies on wild-type and mutant FabIs, and reveal that the complex is primarily stabilized by interactions between acidic residues in the ACP helix alpha2 and a patch of basic residues adjacent to the FabI substrate-binding loop. Unexpectedly, the acyl-pantetheine thioester carbonyl is not hydrogen-bonded to Tyr(156), a conserved component of the short chain alcohol dehydrogenase/reductase superfamily active site triad. FabI is a proven target for drug discovery and the present structure provides insight into the molecular determinants that regulate the interaction of ACPs with target proteins.
[Show abstract][Hide abstract] ABSTRACT: Keeping pace with emerging drug resistance in clinically important pathogens will be greatly aided by inexpensive yet reliable computational methods that predict the binding affinities of ligands for drug targets. We present results using the molecular mechanics Poisson-Boltzmann surface area (MM-PBSA) method to calculate the affinity of a series of triclosan analogues for the E. coli enoyl reductase FabI, spanning a 450000-fold range of binding affinities. Significantly, a high correlation is observed between the calculated binding energies and those determined experimentally. Further examination indicates that the van der Waals energies are the most correlated component of the total affinity (r2 = 0.74), indicating that the shape of the inhibitor is very important in defining the binding energies for this system. The validation of MM-PBSA for the E coli FabI system serves as a platform for inhibitor design efforts focused on the homologous enzyme in Staphylococcus aureus and Mycobacterium tuberculosis.