An investigation into an unusual glycan branching enzyme from Mycobacterium tuberculosis

Mycobacterium tuberculosis (Mtb), the pathogen that causes tuberculosis (TB), is showing increasing resistance to current drug therapies. It is therefore essential that novel drug targets and drug therapies are discovered. Methylglucose lipopolysaccharides (MGLPS) are glycoconjugates produced by Mycobacteria identified as essential for viability. The genes involved in the biosynthesis of MGLPs have been shown to be essential for the survival of the bacterium. Initial stages of the biosynthetic pathway to MGLPs are thought to include the transfer of a glucose unit to glucosyl glycerate (GG), forming di glucosyl glycerate. The gene responsible for this reaction has been identified as Rv3031 and is thought to encode a GH57 glucan branching enzyme (GBE). Known enzymes belonging to this family act upon large substrates, however, the proposed biosynthetic pathway to MGLPs implies that the MtGBE works on much smaller carbohydrate units. The full chemical synthesis of several carbohydrate compounds was performed, including the synthesis of GG and a novel synthetic route to maltosyl glycerate (MalG). Compounds were characterised using NMR spectroscopy and mass spectrometry and GG and MalG were tested in enzymatic assays ... (continues)