Outer Membrane Targeting of <named-content content-type="genus-species">Pseudomonas aeruginosa</named-content> Proteins Shows Variable Dependence on the Components of Bam and Lol Machineries

ABSTRACT In Gram-negative bacteria, the Lol and Bam machineries direct the targeting of lipidated and nonlipidated proteins, respectively, to the outer membrane (OM). Using Pseudomonas aeruginosa strains with depleted levels of specific Bam and Lol proteins, we demonstrated a variable dependence of...

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Autores principales:	Hanh H. Hoang, Nicholas N. Nickerson, Vincent T. Lee, Anastasia Kazimirova, Mohamed Chami, Anthony P. Pugsley, Stephen Lory
Formato:	article
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Publicado:	American Society for Microbiology 2011
Materias:	Microbiology QR1-502
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spelling	oai:doaj.org-article:0f33d4338785498a93ba3cff117275072021-11-15T15:38:48ZOuter Membrane Targeting of <named-content content-type="genus-species">Pseudomonas aeruginosa</named-content> Proteins Shows Variable Dependence on the Components of Bam and Lol Machineries10.1128/mBio.00246-112150-7511https://doaj.org/article/0f33d4338785498a93ba3cff117275072011-12-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mBio.00246-11https://doaj.org/toc/2150-7511ABSTRACT In Gram-negative bacteria, the Lol and Bam machineries direct the targeting of lipidated and nonlipidated proteins, respectively, to the outer membrane (OM). Using Pseudomonas aeruginosa strains with depleted levels of specific Bam and Lol proteins, we demonstrated a variable dependence of different OM proteins on these targeting pathways. Reduction in the level of BamA significantly affected the ability of the β-barrel membrane protein OprF to localize to the OM, while the targeting of three secretins that are functionally related OM proteins was less affected (PilQ and PscC) or not at all affected (XcpQ). Depletion of LolB affected all lipoproteins examined and had a variable effect on the nonlipidated proteins. While the levels of OprF, PilQ, and PscC were significantly reduced by LolB depletion, XcpQ was unaffected and was correctly localized to the OM. These results suggest that certain β-barrel proteins such as OprF primarily utilize the complete Bam machinery. The Lol machinery participates in the OM targeting of secretins to variable degrees, likely through its involvement in the assembly of lipidated Bam components. XcpQ, but not PilQ or PscC, was shown to assemble spontaneously into liposomes as multimers. This work raises the possibility that there is a gradient of utilization of Bam and Lol insertion and targeting machineries. Structural features of individual proteins, including their β-barrel content, may determine the propensity of these proteins for folding (or misfolding) during periplasmic transit and OM insertion, thereby influencing the extent of utilization of the Bam targeting machinery, respectively. IMPORTANCE Targeting of lipidated and nonlipidated proteins to the outer membrane (OM) compartment in Gram-negative bacteria involves the transfer across the periplasm utilizing the Lol and Bam machineries, respectively. We show that depletion of Bam and Lol components in Pseudomonas aeruginosa does not lead to a general OM protein translocation defect, but the severity (and therefore, Lol and Bam dependence), varies with individual proteins. XcpQ, the secretin component of the type II secretion apparatus, is translocated into the OM without the assistance of Bam or Lol machineries. The hypothesis that XcpQ, after secretion across the cytoplasmic membrane, does not utilize the OM targeting machineries was supported by demonstrating that in vitro-synthesized XcpQ (but not the other P. aeruginosa secretins) can spontaneously incorporate into lipid vesicles. Therefore, the requirement for ancillary factors appears to be, in certain instances, dictated by the intrinsic properties of individual OM proteins, conceivably reflecting their propensities to misfold during periplasmic transit.Hanh H. HoangNicholas N. NickersonVincent T. LeeAnastasia KazimirovaMohamed ChamiAnthony P. PugsleyStephen LoryAmerican Society for MicrobiologyarticleMicrobiologyQR1-502ENmBio, Vol 2, Iss 6 (2011)
institution	DOAJ
collection	DOAJ
language	EN
topic	Microbiology QR1-502
spellingShingle	Microbiology QR1-502 Hanh H. Hoang Nicholas N. Nickerson Vincent T. Lee Anastasia Kazimirova Mohamed Chami Anthony P. Pugsley Stephen Lory Outer Membrane Targeting of <named-content content-type="genus-species">Pseudomonas aeruginosa</named-content> Proteins Shows Variable Dependence on the Components of Bam and Lol Machineries
description	ABSTRACT In Gram-negative bacteria, the Lol and Bam machineries direct the targeting of lipidated and nonlipidated proteins, respectively, to the outer membrane (OM). Using Pseudomonas aeruginosa strains with depleted levels of specific Bam and Lol proteins, we demonstrated a variable dependence of different OM proteins on these targeting pathways. Reduction in the level of BamA significantly affected the ability of the β-barrel membrane protein OprF to localize to the OM, while the targeting of three secretins that are functionally related OM proteins was less affected (PilQ and PscC) or not at all affected (XcpQ). Depletion of LolB affected all lipoproteins examined and had a variable effect on the nonlipidated proteins. While the levels of OprF, PilQ, and PscC were significantly reduced by LolB depletion, XcpQ was unaffected and was correctly localized to the OM. These results suggest that certain β-barrel proteins such as OprF primarily utilize the complete Bam machinery. The Lol machinery participates in the OM targeting of secretins to variable degrees, likely through its involvement in the assembly of lipidated Bam components. XcpQ, but not PilQ or PscC, was shown to assemble spontaneously into liposomes as multimers. This work raises the possibility that there is a gradient of utilization of Bam and Lol insertion and targeting machineries. Structural features of individual proteins, including their β-barrel content, may determine the propensity of these proteins for folding (or misfolding) during periplasmic transit and OM insertion, thereby influencing the extent of utilization of the Bam targeting machinery, respectively. IMPORTANCE Targeting of lipidated and nonlipidated proteins to the outer membrane (OM) compartment in Gram-negative bacteria involves the transfer across the periplasm utilizing the Lol and Bam machineries, respectively. We show that depletion of Bam and Lol components in Pseudomonas aeruginosa does not lead to a general OM protein translocation defect, but the severity (and therefore, Lol and Bam dependence), varies with individual proteins. XcpQ, the secretin component of the type II secretion apparatus, is translocated into the OM without the assistance of Bam or Lol machineries. The hypothesis that XcpQ, after secretion across the cytoplasmic membrane, does not utilize the OM targeting machineries was supported by demonstrating that in vitro-synthesized XcpQ (but not the other P. aeruginosa secretins) can spontaneously incorporate into lipid vesicles. Therefore, the requirement for ancillary factors appears to be, in certain instances, dictated by the intrinsic properties of individual OM proteins, conceivably reflecting their propensities to misfold during periplasmic transit.
format	article
author	Hanh H. Hoang Nicholas N. Nickerson Vincent T. Lee Anastasia Kazimirova Mohamed Chami Anthony P. Pugsley Stephen Lory
author_facet	Hanh H. Hoang Nicholas N. Nickerson Vincent T. Lee Anastasia Kazimirova Mohamed Chami Anthony P. Pugsley Stephen Lory
author_sort	Hanh H. Hoang
title	Outer Membrane Targeting of <named-content content-type="genus-species">Pseudomonas aeruginosa</named-content> Proteins Shows Variable Dependence on the Components of Bam and Lol Machineries
title_short	Outer Membrane Targeting of <named-content content-type="genus-species">Pseudomonas aeruginosa</named-content> Proteins Shows Variable Dependence on the Components of Bam and Lol Machineries
title_full	Outer Membrane Targeting of <named-content content-type="genus-species">Pseudomonas aeruginosa</named-content> Proteins Shows Variable Dependence on the Components of Bam and Lol Machineries
title_fullStr	Outer Membrane Targeting of <named-content content-type="genus-species">Pseudomonas aeruginosa</named-content> Proteins Shows Variable Dependence on the Components of Bam and Lol Machineries
title_full_unstemmed	Outer Membrane Targeting of <named-content content-type="genus-species">Pseudomonas aeruginosa</named-content> Proteins Shows Variable Dependence on the Components of Bam and Lol Machineries
title_sort	outer membrane targeting of <named-content content-type="genus-species">pseudomonas aeruginosa</named-content> proteins shows variable dependence on the components of bam and lol machineries
publisher	American Society for Microbiology
publishDate	2011
url	https://doaj.org/article/0f33d4338785498a93ba3cff11727507
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Outer Membrane Targeting of <named-content content-type="genus-species">Pseudomonas aeruginosa</named-content> Proteins Shows Variable Dependence on the Components of Bam and Lol Machineries

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