The results of NGS were compared with those obtained by conventional SS

chanistic and structural information on the target enzyme. Enzymes offer unique opportunities for drug design that are not available to cell surface receptors, nuclear hormone receptors, ion channel, transporters, and DNA. It has been pointed out that one of the lessons to be learned from marketed enzyme inhibitors is that the most potent and effective inhibitors take advantage of enzyme chemistry to achieve inhibition. Moreover, the recognition of the limitations M. tuberculosis Shikimate Kinase of high-throughput screening approaches in the discovery of candidate drugs has rekindled interest in rational design methods. Accordingly, mechanistic analysis should always be a top priority for enzyme-targeted drug programs aiming at the rational design of potent enzyme inhibitors. Moreover, targets that are both essential to survival of, and exclusive to, M. tuberculosis are particularly promising as their inhibition could lead to the development of non-toxic drugs to the human host and having effective killing effect on the pathogen. The biosynthesis of aromatic rings from carbohydrate precursors involves a range of chemical transformations that together constitute the shikimate pathway; through seven enzymatic steps, phosphoenolpyruvate and D-erythrose 4-phosphate are condensed to the branch point compound chorismate, which leads to several additional terminal pathways. The shikimate pathway is essential in algae, higher plants, fungi, bacteria, apicomplexan parasites and sea anemone, but absent from humans. The mycobacterial shikimate pathway leads to the 7623957 biosynthesis of chorismic acid, which is converted by five distinct enzymes to prephenate, anthranilate, aminodeoxychorismate, para-hydroxybenzoic acid, and isochorismate . The aroK-encoded M. tuberculosis Shikimate Kinase, the fifth enzyme of the pathway, catalyzes a phosphoryl transfer from ATP to the carbon-3 hydroxyl group of shikimate ]3,4,5-trihydroxy-1-cyclohexene-1-carboxylic acid) forming shikimate 3-phosphate . Disruption of aroK gene has demonstrated that MtSK, and thus the common aromatic biosynthesis pathway, is essential for the viability of M. tuberculosis. We have previously reported cloning and expression in Escherichia coli of recombinant MtSK in functional form, thereby confirming the correct in silico assignment to the structural gene encoding this protein. Our research group and others have reported crystal structure determinations of MtSK. Three functional motifs of nucleotide-binding enzymes were recognizable in MtSK, including a Walker A-motif, a Walker B-motif, and an adenine-binding loop. MtSK belongs to the family of nucleoside monophosphate kinases, which are composed of three domains: the CORE domain containing the five stranded MedChemExpress SNDX 275 parallel b-sheet and the P-loop, which forms the binding site for nucleotides; the LID domain, which closes over the active site and has residues that are essential for the binding of ATP; and the NMP-binding domain, which functions to recognize and bind shikimate. More recently, based on an analysis of global movements upon ligand binding, it has 22884612 been proposed that MtSK is comprised of four domains: the ESB domain; the nucleotide-binding site that includes the P-loop, the ABloop, and the segment of 101110; the LID domain; and the Reduced Core domain. A characteristic feature of NMP kinases is that they undergo large conformational changes during catalysis. Based on a series of high-resolution crystal structures of MtSK in apo form a