This Article Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Reprints and Permissions Copyright Information Books from ASM Press MicrobeWorld Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Gabay, J E Articles by Horwitz, M A Search for Related Content PubMed PubMed Citation Articles by Gabay, J E Articles by Horwitz, M A

Purification of Legionella pneumophila major outer membrane protein and demonstration that it is a porin.

ABSTRACT

We have purified the major outer membrane protein (MOMP) of Legionella pneumophila, determined that it is associated with peptidoglycan, and characterized it as a porin. To purify the MOMP, we used a simple, rapid, three-step procedure that gave us the protein in high yield. The first step of the purification procedure involved selectively extracting the MOMP from whole bacterial cells with calcium and zwitterionic detergent. The second and third steps achieved purification by ion-exchange and molecular-sieve chromatography. The dissociation of the MOMP into monomers was dependent upon the presence of a reducing agent and was enhanced by treatment at 100 degrees C. To study the relationship of the MOMP to peptidoglycan, we extracted the protein by a modification of the Rosenbusch procedure. Like the Escherichia coli porins, the MOMP was peptidoglycan associated. The MOMP was at least partially dissociated from peptidoglycan in sodium dodecyl sulfate and a high salt concentration. To study the ion channel-forming properties of the MOMP, we reconstituted the MOMP in planar lipid membranes. The MOMP formed ion-permeable channels with a single-channel conductance size of 100 picoSiemens. The MOMP channels exhibited a fourfold selectivity for cations over anions and voltage-independent gating. These findings demonstrate that the MOMP is a porin with properties similar to those of E. coli porins.

This article has been cited by other articles:

• Losick, V. P., Isberg, R. R. (2006). NF-{kappa}B translocation prevents host cell death after low-dose challenge by Legionella pneumophila. J. Exp. Med. 203: 2177-2189 [Abstract] [Full Text]  
• VanRheenen, S. M., Luo, Z.-Q., O'Connor, T., Isberg, R. R. (2006). Members of a Legionella pneumophila Family of Proteins with ExoU (Phospholipase A) Active Sites Are Translocated to Target Cells.. Infect. Immun. 74: 3597-3606 [Abstract] [Full Text]  
• Derre, I., Isberg, R. R. (2005). LidA, a Translocated Substrate of the Legionella pneumophila Type IV Secretion System, Interferes with the Early Secretory Pathway. Infect. Immun. 73: 4370-4380 [Abstract] [Full Text]  
• Derre, I., Isberg, R. R. (2004). Macrophages from Mice with the Restrictive Lgn1 Allele Exhibit Multifactorial Resistance to Legionella pneumophila. Infect. Immun. 72: 6221-6229 [Abstract] [Full Text]  
• VanRheenen, S. M., Dumenil, G., Isberg, R. R. (2004). IcmF and DotU Are Required for Optimal Effector Translocation and Trafficking of the Legionella pneumophila Vacuole. Infect. Immun. 72: 5972-5982 [Abstract] [Full Text]  
• Nikaido, H. (2003). Molecular Basis of Bacterial Outer Membrane Permeability Revisited. Microbiol. Mol. Biol. Rev. 67: 593-656 [Abstract] [Full Text]  
• Rossier, O., Cianciotto, N. P. (2001). Type II Protein Secretion Is a Subset of the PilD-Dependent Processes That Facilitate Intracellular Infection by Legionella pneumophila. Infect. Immun. 69: 2092-2098 [Abstract] [Full Text]  
• Viswanathan, V. K., Edelstein, P. H., Pope, C. D., Cianciotto, N. P. (2000). The Legionella pneumophila iraAB Locus Is Required for Iron Assimilation, Intracellular Infection, and Virulence. Infect. Immun. 68: 1069-1079 [Abstract] [Full Text]  
• Shannon, J. L., Fernandez, R. C. (1999). The C-Terminal Domain of the Bordetella pertussis Autotransporter BrkA Forms a Pore in Lipid Bilayer Membranes. J. Bacteriol. 181: 5838-5842 [Abstract] [Full Text]