Published in
Public Library of Science, PLoS Pathogens, 3(6), p. e1000794, 2010
DOI: 10.1371/journal.ppat.1000794
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Tools
Direct Visualization by Cryo-EM of the Mycobacterial Capsular Layer: A Labile Structure Containing ESX-1-Secreted Proteins
,
Jeroen Geurtsen,
Jason Pierson,
Karin de Punder,
Maaike van Zon,
Brigitte Wever,
Sander R. Piersma,
Connie R. Jiménez,
Mamadou Daffé,
Ben J. Appelmelk,
Wilbert Bitter ,
Nicole van der Wel ,
Peter J. Peters
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Abstract
The cell envelope of mycobacteria, a group of Gram positive bacteria, is composed of a plasma membrane and a Gram-negative-like outer membrane containing mycolic acids. In addition, the surface of the mycobacteria is coated with an ill-characterized layer of extractable, non-covalently linked glycans, lipids and proteins, collectively known as the capsule, whose occurrence is a matter of debate. By using plunge freezing cryo-electron microscopy technique, we were able to show that pathogenic mycobacteria produce a thick capsule, only present when the cells were grown under unperturbed conditions and easily removed by mild detergents. This detergent-labile capsule layer contains arabinomannan, alpha-glucan and oligomannosyl-capped glycolipids. Further immunogenic and proteomic analyses revealed that Mycobacterium marinum capsule contains high amounts of proteins that are secreted via the ESX-1 pathway. Finally, cell infection experiments demonstrated the importance of the capsule for binding to cells and dampening of pro-inflammatory cytokine response. Together, these results show a direct visualization of the mycobacterial capsular layer as a labile structure that contains ESX-1-secreted proteins.