Cryo-EM Structures and Regulation of Arabinofuranosyltransferase AftD from Mycobacteria

Tan, Y. Z.; Zhang, L.; Rodrigues, J.; Zheng, R. B.; Giacometti, S. I.; Rosario, A. L.; Kloss, B.; Dandey, V. P.; Wei, H.; Brunton, R.; Raczkowski, A. M.; Athayde, D.; Catalao, M. J.; Pimentel, M.; Clarke, O. B.; Lowary, T. L.; Archer, M.; Niederweis, M.; Potter, C. S.; Carragher, B.; Mancia, F.

doi:10.1101/2019.12.22.885152

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bioRxiv · 10.1101/2019.12.22.885152

Cryo-EM Structures and Regulation of Arabinofuranosyltransferase AftD from Mycobacteria

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Abstract

Mycobacterium tuberculosis causes tuberculosis, a disease that kills over one million people each year. Its cell envelope is a common antibiotic target and has a unique structure due, in part, to two lipidated polysaccharides - arabinogalactan and lipoarabinomannan. Arabinofuranosyltransferase D (AftD) is an essential enzyme involved in assembling these glycolipids. We present the 2.9 [A] resolution structure of M. abscessus AftD determined by single particle cryo-electron microscopy. AftD has a conserved GT-C glycosyltransferase fold and three carbohydrate binding modules. Glycan array analysis shows that AftD binds complex arabinose glycans. Additionally, AftD is non-covalently complexed with an acyl carrier protein (ACP). 3.4 and 3.5 [A] structures of a mutant with impaired ACP binding reveal a conformational change that suggests the ACP may regulate AftD function. Using a conditional knock-out constructed in M. smegmatis, mutagenesis experiments confirm the essentiality of the putative active site and the ACP binding for AftD function.

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Tan, Y. Z., Zhang, L., Rodrigues, J., Zheng, R. B., Giacometti, S. I., Rosario, A. L., Kloss, B., Dandey, V. P., Wei, H., Brunton, R., Raczkowski, A. M., Athayde, D., Catalao, M. J., Pimentel, M., Clarke, O. B., Lowary, T. L., Archer, M., Niederweis, M., Potter, C. S., Carragher, B., Mancia, F.. 2019-12-23. Cryo-EM Structures and Regulation of Arabinofuranosyltransferase AftD from Mycobacteria. https://doi.org/10.1101/2019.12.22.885152

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