1. Mycobacterium tuberculosis chaperonin 10 is secreted in the macrophage phagosome: Is secretion due to dissociation and adoption of a partially helical structure at the membrane?
- Author
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Elena Giannozzi, Francesca Mangili, Emanuela Gancia, Neri Niccolai, Paolo Mascagni, Anthony R.M. Coates, Emanuele Rizzi, Gianluca Fossati, Maria Luisa Moras, Michael M. Roberts, Brian Henderson, Gaetano Izzo, Daniela Modena, L. Bono, Christopher Walters, Eugenio Biagio Leone, Ottavia Spiga, Stephen E. Harding, Neil Errington, Piero Marone, Bruno Casetta, Fossati, G, Izzo, G, Rizzi, E, Gancia, E, Modena, D, Moras, M, Niccolai, N, Giannozzi, E, Spiga, O, Bono, L, Marone, P, Leone, B, Mangili, F, Harding, S, Errington, N, Walters, C, Henderson, B, Roberts, M, Coates, A, Casetta, B, and Mascagni, P
- Subjects
Circular dichroism ,Magnetic Resonance Spectroscopy ,Biology ,Microbiology ,Mass Spectrometry ,Protein Structure, Secondary ,Chaperonin ,Cell Line ,Cell membrane ,Mice ,Structure-Activity Relationship ,Protein structure ,Structural Biology ,Phagosomes ,medicine ,Chaperonin 10 ,Animals ,Humans ,Animals, Cell Line, Cell Membrane/metabolism, Chaperonin 10/chemistry, Chaperonin 10/genetics, Chaperonin 10/metabolism, Circular Dichroism Humans, Macrophages/metabolism, Macrophages/microbiology, Magnetic Resonance Spectroscopy, Mass Spectrometry/methods, Mice, Microscopy Electron, Mycobacterium tuberculosis/metabolism, Mycobacterium tuberculosis/pathogenicity, Peptides/chemical synthesis, Peptides/chemistry, Phagosomes/metabolism, Phagosomes/microbiology, Protein Structure, Secondary Rabbits, Solvents, Structure-Activity Relationship ,Molecular Biology ,Protein secondary structure ,Phagosome ,Circular Dichroism ,Macrophages ,Cell Membrane ,Biological membrane ,GroES ,Mycobacterium tuberculosis ,Microscopy, Electron ,medicine.anatomical_structure ,Biochemistry ,MED/07 - MICROBIOLOGIA E MICROBIOLOGIA CLINICA ,Solvents ,Rabbits ,Peptides - Abstract
To confirm thatMycobacterium tuberculosischaperonin 10 (Cpn10) is secreted outside the live bacillus, infected macrophages were examined by electron microscopy. This revealed that the mycobacterial protein accumulates both in the wall of the bacterium and in the matrix of the phagosomes in which ingested mycobacteria survive within infected macrophages. To understand the structural implications underlying this secretion, a structural study ofM.tuberculosisCpn10 was performed under conditions that are generally believed to mimic the membrane environment. It was found that in buffer-organic solvent mixtures, the mycobacterial protein forms two main species, namely, a partially helical monomer that prevails in dilute solutions at room temperature and a dimer that folds into a β-sheet-dominated structure and prevails in either concentrated protein solutions at room temperature or in dilute solutions at low temperature. A partially helical monomer was also found and was completely associated with negatively charged detergents in a micelle-bound state. Remarkably, zwitterionic lipids had no effect on the protein structure. By using N- and C-truncated forms of the protein, the C- and N-terminal sequences were identified as possessing an amphiphilic helical character and as selectively associating with acidic detergent micelles. When the study was extended to other chaperonins, it was found that human Cpn10 is also monomeric and partially helical in dilute organic solvent-buffer mixtures. In contrast,Escherichia coliCpn10 is mostly dimeric and predominately β-sheet in both dilute and concentrated solutions. Interestingly, human Cpn10 also crosses biological membranes, whereas theE.colihomologue is strictly cytosolic. These results suggest that dissociation to partially helical monomers and interaction with acidic lipids may be two important steps in the mechanism of secretion ofM.tuberculosisCpn10 to the external environment.
- Published
- 2003