Mycobacterial arabinan biosynthesis: the use of synthetic arabinoside acceptors in the development of an arabinosyl transfer assay |
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Authors: | Lee, Richard E. Brennan, Patrick J. Besra, Gurdyal S. |
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Affiliation: | Department of Microbiology, Colorado State University Fort Collins, CO 80523-1677, USA |
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Abstract: | Information on the biosynthesis of the D-arabinans of the cellwall of Mycobacterium tuberculosis is rapidly emerging, withthe promise of new targets for drug development against tuberculosis.Accordingly, arabinosyl transferase assays were developed utilizingsynthesized [1–14C]-β-D-arabinofuranosyl-1-monophosphoryldecaprenolas donor and a variety of O- and S-alkyl arabinosides as acceptors.These were: -D-Araf-(15)--D-Araf-O- and -S-alkyl di-arabinosidesand -D-Araf-(15)--D-Araf-(15)--D-Araf-O- and -S-alkyl triarabinosides.Whereas the O- and S-alkyl monosaccharide acceptors were inactive,the O- and S-alkyl disaccharide and the O- and S-alkyl trisaccharideacceptors (O-alkyl disaccharide acceptors with a C8alkyl chain were more active than those containing the C6 orC10 analogs. Chemical analysis of the enzymatically synthesizedproducts of the reactions demonstrated that β-D-arabinofuranosyl-1-monophosphoryldecaprenolwas an effective donor for two of the three potential arabinosyltransferases: β-D-arabinofuranosyl-1-monophosphoryldecaprenol:arabinan (15) arabinosyl transferase and β-D-arabinofuranosyl-1-monophosphoryl-decaprenol:arabinan β(12) arabinosyl transferase. The β(12) arabinosyltransferase activity was more in evidence in the presence ofthe O-alkyl disaccharide acceptor, whereas both transferaseswere about equivalent in the presence of the S-alkyl trisaccharideacceptor. The tuberculosis drug, ethambutol, a known mycobacterialarabinosyl transferase inhibitor, was inactive within thesearabinosyl transferase/acceptor based assay systems, supportingother evidence that a third activity, responsible for the formationof 13 linkage, is the drug target. acceptor arabinan biosynthesis glycosyltrans-ferase assay mycobacteria |
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