One-Pot Phosphate-Mediated Synthesis of Novel 1,3,5-Trisubstituted Pyridinium Salts: A New Family of S. aureus Inhibitors

Polysubstituted pyridinium salts are valuable pharmacophores found in many biologically active molecules. Their synthesis typically involves the use of multistep procedures or harsh reaction conditions. Here, we report water-based phosphate mediated reaction conditions that promote the condensation of arylacetaldehydes with amines to give 1,3,5-pyridinium salts. The reaction, carried out at pH 6, provides conditions suitable for the use of less stable aldehydes and amines in this Chichibabin pyridine condensation. The evaluation of selected 1,3,5-trisubstituted pyridinium salts highlighted that they can inhibit the growth of S. aureus in the low μg/mL range. The synthetic accessibility of these compounds and preliminary growth inhibition data may pave the way towards the discovery of new anti-bacterials based on the 1,3,5-trisubstituted pyridinium scaffold

'Dopamine-first' mechanism enables the rational engineering of the norcoclaurine synthase aldehyde activity profile

Norcoclaurine synthase (NCS) (EC 4.2.1.78) catalyzes the Pictet–Spengler condensation of dopamine and an aldehyde, forming a substituted (S)-tetrahydroisoquinoline, a pharmaceutically important moiety. This unique activity has led to NCS being used for both in vitro biocatalysis and in vivo recombinant metabolism. Future engineering of NCS activity to enable the synthesis of diverse tetrahydroisoquinolines is dependent on an understanding of the NCS mechanism and kinetics. We assess two proposed mechanisms for NCS activity: (a) one based on the holo X-ray crystal structure and (b) the ‘dopamine-first’ mechanism based on computational docking. Thalictrum flavum NCS variant activities support the dopamine-first mechanism. Suppression of the non-enzymatic background reaction reveals novel kinetic parameters for NCS, showing it to act with low catalytic efficiency. This kinetic behaviour can account for the ineffectiveness of recombinant NCS in in vivo systems, and also suggests NCS may have an in planta role as a metabolic gatekeeper. The amino acid substitution L76A, situated in the proposed aldehyde binding site, results in the alteration of the enzyme's aldehyde activity profile. This both verifies the dopamine-first mechanism and demonstrates the potential for the rational engineering of NCS activity

One-pot triangular chemoenzymatic cascades for the syntheses of chiral alkaloids from dopamine

We describe novel chemoenzymatic routes to (S)-benzylisoquinoline and (S)-tetrahydroprotoberberine alkaloids using the enzymes transaminase (TAm) and norcoclaurine synthase (NCS) in a one-pot, one-substrate ‘triangular’ cascade. Employment of up to two C–C bond forming steps allows for the rapid generation of molecular complexity under mild conditions

Tetrahydroisoquinolines affect the whole-cell phenotype of Mycobacterium tuberculosis by inhibiting the ATP-dependent MurE ligase

Objectives (S)-Leucoxine, isolated from the Colombian Lauraceae tree Rhodostemonodaphne crenaticupula Madriñan, was found to inhibit the growth of Mycobacterium tuberculosis H37Rv. A biomimetic approach for the chemical synthesis of a wide array of 1-substituted tetrahydroisoquinolines was undertaken with the aim of elucidating a common pharmacophore for these compounds with novel mode(s) of anti-TB action. Methods Biomimetic Pictet–Spengler or Bischler–Napieralski synthetic routes were employed followed by an evaluation of the biological activity of the synthesized compounds. Results In this work, the synthesized tetrahydroisoquinolines were found to inhibit the growth of M. tuberculosis H37Rv and affect its whole-cell phenotype as well as the activity of the ATP-dependent MurE ligase, a key enzyme involved in the early stage of cell wall peptidoglycan biosynthesis. Conclusions As the correlation between the MIC and the half-inhibitory enzymatic concentration was not particularly strong, there is a credible possibility that these compounds have pleiotropic mechanism(s) of action in M. tuberculosis

Discovery of Inhibitors of Leishmania β-1,2-Mannosyltransferases Using a Click-Chemistry-Derived Guanosine Monophosphate Library

Leishmania spp. are a medically important group of protozoan parasites that synthesize a novel intracellular carbohydrate reserve polymer termed mannogen. Mannogen is a soluble homopolymer of β-1,2-linked mannose residues that accumulates in the major pathogenic stages in the sandfly vector and mammalian host. While several steps in mannogen biosynthesis have been defined, none of the enzymes have been isolated or characterized. We report the development of a simple assay for the GDP-mannose–dependent β-1,2-mannosyltransferases involved in mannogen synthesis. This assay utilizes octyl α-d-mannopyranoside to prime the formation of short mannogen oligomers up to 5 mannose residues. This assay was used to screen a focussed library of 44 GMP-triazole adducts for inhibitors. Several compounds provided effective inhibition of mannogen β-1,2-mannosyltransferases in a cell-free membrane preparation. This assay and inhibitor compounds will be useful for dissecting the role of different mannosyltransferases in regulating de novo biosynthesis and elongation reactions in mannogen metabolism

A survey of chemical methods for sugar-nucleotide synthesis

Covering: up to 2009. Sugar-nucleotides are essential intermediates in carbohydrate metabolism and glycoconjugate biosynthesis. Structurally, sugar-nucleotides are composed of a sugar or sugar derivative and a nucleoside mono- or diphosphate. Numerous combinations of sugars and nucleotides are found in nature, and although only nine different sugar-nucleotides have so far been identified in mammalian cells, this number is much greater in other organisms. Naturally occurring sugar-nucleotides, as well as structural analogues, are of great interest as substrates for enzymatic reactions in carbohydrate synthesis, as enzyme inhibitors, as tools for assay development and for the study of glycoconjugate biosynthesis. Therefore, methods for the efficient preparation of natural and non-natural sugar-nucleotides are of considerable importance for synthetic, biological and medicinal chemistry. The synthesis of sugar-nucleotides is non-trivial and complicated by a number of factors: the low solubility of sugar-nucleotides in organic solvents, the presence of several polar or charged functional groups, and the susceptibility of the glycosidic and pyrophosphate bonds to hydrolytic cleavage. To address these issues, both chemical and enzymatic strategies have been pursued. Here we present an overview of current methods for the chemical synthesis of NDP- and NMP-sugars, including recently developed protocols for the direct chemical manipulation of unprotected sugar-nucleotides

Structural and mechanistic basis for a new mode of glycosyltransferase inhibition

Glycosyltransferases are carbohydrate-active enzymes with essential roles in numerous important biological processes. We have developed a novel donor analogue for galactosyltransferases which locks a representative target enzyme in a catalytically inactive conformation, thus almost completely abolishing sugar transfer. Results with other galactosyltransferases suggest that this novel and unique mode of glycosyltransferase inhibition is, very likely, generally applicable to other members of this very important enzyme family also