Design, Synthesis, and X-ray Analysis of a Glycoconjugate Bound to Mycobacterium tuberculosis Antigen 85C

Abstract

Tuberculosis (TB) is a global health threat with nearly 500,000 new cases of multidrug-resistance TB estimated to occur every year, so new drugs are desperately needed. A number of current antimycobacterial drugs work by interfering with the biosynthesis of key components of the mycolylarabinogalactan (mAG). In light of this observation, other enzymes involved in the synthesis of the mAG should also serve as targets for antimycobacterial drug development. One potential target is the Antigen 85 (Ag85) complex, a family of mycolyltransferases that are responsible for the transfer of mycolic acids from trehalose monomycolate (TMM) to the arabinogalactan. Virtual thiophenyl-arbinoside conjugates were docked to antigen Ag85C (PDB code: 1va5) using Glide. Compounds with good docking scores were synthesized by a Gewald synthesis followed by linking to 5-thioarabinofuranosides. The resulting thiophenyl-thioarabinofuranosides were assayed for inhibition of mycoyltransferase activity using a 4-methylumbelliferyl butyrate fluorescence assay. The conjugates showed Ki values ranging from 18.2 to 71.0 µM. The most potent inhibitor was soaked into crystals of Mycobacterium tuberculosis antigen 85C and the structure of the complex determined. The X-ray structure shows the compound bound within the active site of the enzyme with the thiophene moiety is positioned in the putative α-chain binding site of TMM and the arabinofuranoside moiety within the known carbohydrate-binding site as exhibited for the Ag85B-trehalose crystal structure. Unexpectedly, no specific hydrogen bonding interactions are being formed between the arabinofuranoside and the carbohydrate-binding site of the active site suggesting that the binding of the arabinoside within this structure is driven by shape complementarily between the arabinosyl moiety and the carbohydrate binding site.

The molecular biology of mycobacterial trehalose in the quest for advanced tuberculosis therapies

“…These enzymes catalyse the transesterification of the lipid moiety of TMM to another TMM molecule to yield TDM, or to AG to assemble the mAPG complex (Armitige et al, 2000;Belisle et al, 1997;Ibrahim et al, 2012;Kaur et al, 2009). In this process, a trehalose molecule is released and may be recycled back into the cytoplasm by the specific LpqY-SugABC transporter system identified recently (see above).…”

New Approaches to Target the Mycolic Acid Biosynthesis Pathway for the Development of Tuberculosis Therapeutics

“…Ibrahim and colleagues developed a series of glycoconjugates based on the fragments above as potential inhibitors of the Ag85 complex [148]. They tested these compounds against Ag85C in the presence of 4-methylumbelliferyl butyrate (4MUB), a synthetic substrate that when Ag85C binds 4MUB, the fluorescent 4-methylumbelliferone is released [148].…”

“…They tested these compounds against Ag85C in the presence of 4-methylumbelliferyl butyrate (4MUB), a synthetic substrate that when Ag85C binds 4MUB, the fluorescent 4-methylumbelliferone is released [148]. Therefore, suppression of a fluorescent signal corresponds to Ag85C inhibition.…”

Synthesis of a Poly-hydroxypyrolidine-Based inhibitor of Mycobacterium tuberculosis GlgE

“…2 Following the mycolyl transfer reaction, trehalose is transferred back into the cytoplasm from the periplamic space and is modified in the production of various trehalolipids and sulfolipids. 3,4 Trehalose is also converted into a branched α-glucan. The conversion process involves four enzymes: trehalose synthase (TreS), maltokinase (Pep2), GlgE, and GlgB.…”