Chaperone Spy Protects Outer Membrane Proteins from Folding Stress via Dynamic Complex Formation

He, Wei; Yu, Gangjin; Li, Tianpeng; Bai, Lijun; Yang, Yuanyuan; Xue, Zixiao; Pang, Yonghao; Reichmann, Dana; Hiller, Sebastian; He, Lichun; Liu, Maili; Qin, Shu

doi:10.1128/mbio.02130-21

Cited by 17 publications

(12 citation statements)

References 58 publications

(83 reference statements)

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“…Studies had confirmed that antibacterial drugs were blocked from the target by Omp channels (Omp could restrict the entry of antibacterial drugs into cells by controlling the pore size of the channel) before they entered the bacteria, and it could also form an efflux system to remove the antibacterial drugs that had entered the cell. It was pumped out of the cell, but some antibacterial drugs could achieve an antibacterial effect by increasing the permeability of Omp [ 23 , 24 ]. When a certain Omp function was inhibited, the bacteria would activate the expression of other Omp for their own survival.…”

Section: Discussionmentioning

confidence: 99%

Effects of Cinnamon Essential Oil on Oxidative Damage and Outer Membrane Protein Genes of Salmonella enteritidis Cells

Zhang

Zhao

Chen

et al. 2022

Foods

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Salmonella is an important pathogen causing food poisoning. Food safety and health are the themes of today′s society. As a class of food-borne pathogens, Salmonella enteritidis had become one of the common zoonotic pathogens. Cinnamon essential oil (CEO) had been reported as an antibacterial agent, but there are few studies on its antibacterial mechanism. This study investigated the effects of CEO on oxidative damage and outer membrane protein genes of Salmonella enteritidis cells. First, the reactive oxygen species content in bacteria treated with different concentrations of cinnamon essential oil was determined by fluorescence spectrophotometry, and the effects of superoxide dismutase (SOD), catalase (CAT) and superoxide dismutase (SOD), and catalase (CAT) and peroxidase (POD) were determined by the kit method. The activity of POD and the content of malondialdehyde (MDA) were investigated to investigate the oxidative damage of CEO to Salmonella enteritidis cells. By analyzing the effect of CEO on the Salmonella enteritidis cell membrane’s outer membrane protein gene expression, the mechanism of CEO′s action on the Salmonella enteritidis cell membrane was preliminarily discussed. The results showed that CEO treatment had an obvious oxidative damaging effect on Salmonella enteritidis. Compared with the control group, the increase in CEO concentration caused a significant increase in the bacteria ROS content. The observation technique experiment found that with the increase in CEO concentration, the number of stained cells increased, which indicated that CEO treatment would increase the ROS level in the cells, and it would also increase with the increase in CEO concentration, thus causing the oxidation of cells and damage. In addition, CEO treatment also caused the disruption of the balance of the cellular antioxidant enzymes (SOD, CAT, POD) system, resulting in an increase in the content of MDA, a membrane lipid metabolite, and increased protein carbonylation, which ultimately inhibited the growth of Salmonella enteritidis. The measurement results of cell membrane protein gene expression levels showed that the Omp genes to be detected in Salmonella enteritidis were all positive, which indicated that Salmonella enteritidis carried these four genes. Compared with the control group, the relative expressions of OmpF, OmpA and OmpX in the CEO treatment group were significantly increased (p < 0.05), which proved that the cell function was disturbed. Therefore, the toxicity of CEO to Salmonella enteritidis could be attributed to the damage of the cell membrane and the induction of oxidative stress at the same time. It was speculated that the antibacterial mechanism of CEO was the result of multiple effects. This work was expected to provide a theoretical basis for the development of new natural food preservatives and the prevention and control of Salmonella enteritidis.

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Section: Discussionmentioning

confidence: 99%

Effects of Cinnamon Essential Oil on Oxidative Damage and Outer Membrane Protein Genes of Salmonella enteritidis Cells

Zhang

Zhao

Chen

et al. 2022

Foods

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“…OMP biogenesis requires a network of chaperones to keep uOMPs in an unfolded but folding-competent state in the aqueous environment of the periplasm. SurA, Skp, DegP, Spy, and FkpA have all been implicated in this periplasmic chaperone network, 8,9 but the function of FkpA with respect to the OMP biogenesis pathway has been less investigated as compared to other chaperones (Figure 7). Here we show that FkpA increases the folded population of OmpA 171 into diC 11 PC LUVs after one hour but decreases the observed rate of OmpA 171 folding.…”

Section: Discussionmentioning

confidence: 99%

“…Major players in the OMP biogenesis chaperone network include SurA, Skp, and FkpA with additional contributions from Spy and the protease chaperone DegP (Figure 1). 8,9 While much of the recent work elucidating the mechanisms of these ATP-independent chaperones focuses on the chaperones SurA and Skp, 10,11,20,21,[12][13][14][15][16][17][18][19] thermodynamic and mechanistic . CC-BY-NC-ND 4.0 International license available under a (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity.…”

Section: Introductionmentioning

confidence: 99%

“…Major players in the OMP biogenesis chaperone network include SurA, Skp, and FkpA with additional contributions from Spy and the protease chaperone DegP (Figure 1). 8,9 While much of the recent work elucidating the mechanisms of these ATP-independent chaperones focuses on the chaperones SurA and Skp, 10,11,20,21,[12][13][14][15][16][17][18][19] thermodynamic and mechanistic 6 information about the function of FkpA in OMP biogenesis remains sparse. 22,23 Structurally, FkpA forms homodimers and comprises two structural domains: an N-terminal, α-helical domain that mediates dimerization and a C-terminal domain homologous to FK506-binding proteins and capable of catalyzing peptidyl-prolyl isomerization reactions.…”

Section: Introductionmentioning

confidence: 99%

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FkpA Enhances Membrane Protein Folding using an Extensive Interaction Surface

Devlin

Marx

Roskopf

et al. 2022

Preprint

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Outer membrane protein (OMP) biogenesis in gram-negative bacteria is managed by a network of periplasmic chaperones that includes SurA, Skp, and FkpA. These chaperones bind unfolded OMPs (uOMPs) in dynamic conformational ensembles to suppress uOMP aggregation, facilitate diffusion across the periplasm, and enhance OMP folding. FkpA primarily responds to heat-shock stress, but its mechanism is comparatively understudied. To determine FkpA chaperone function, we monitored the folding of a cognate client uOmpA171and found that FkpA increases the folded uOmpA171population but also slows the folding rate, dual functions distinct from the other periplasmic chaperones. The results indicate that FkpA behaves as a chaperone and not as a folding catalyst to directly influence the uOmpA171folding trajectory. We determine the binding affinity between FkpA and uOmpA171by globally fitting sedimentation velocity titrations and found it to be intermediate between the known affinities of Skp and SurA for uOMP clients. Notably, complex formation steeply depends on the urea concentration, suggestive of an extensive binding interface. Initial characterizations of the complex using photo-crosslinking indicates that the binding interface spans the inner surfaces of the entire FkpA molecule. In contrast to prior findings, folding and binding experiments performed using subdomain constructs of FkpA demonstrate that the full-length chaperone is required for full activity. Together these results support that FkpA has a distinct and direct effect on uOMP folding and that it achieves this by utilizing an extensive chaperone-client interface.

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“…Major players in the OMP biogenesis chaperone network include SurA, Skp, and FkpA with additional contributions from Spy and the protease chaperone DegP (Figure 1) (He et al, 2021; Missiakas et al, 1996). While much of the recent work elucidating the mechanisms of these ATP‐independent chaperones focuses on the chaperones SurA and Skp (Burmann et al, 2013; Calabrese et al, 2020; Holdbrook et al, 2017; Humes et al, 2019; Jia et al, 2020; Marx et al, 2020a, 2020b; Mas et al, 2020; Pan et al, 2020; Sandlin et al, 2015; Schiffrin et al, 2017; Thoma et al, 2015), thermodynamic and mechanistic information about the function of FkpA in OMP biogenesis remains sparse (Ge et al, 2014; Ruiz‐Perez et al, 2010).…”

Section: Introductionmentioning

confidence: 99%

FkpA enhances membrane protein folding using an extensive interaction surface

et al. 2023

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Outer membrane protein (OMP) biogenesis in gram‐negative bacteria is managed by a network of periplasmic chaperones that includes SurA, Skp, and FkpA. These chaperones bind unfolded OMPs (uOMPs) in dynamic conformational ensembles to suppress aggregation, facilitate diffusion across the periplasm, and enhance folding. FkpA primarily responds to heat‐shock stress, but its mechanism is comparatively understudied. To determine FkpA chaperone function in the context of OMP folding, we monitored the folding of three OMPs and found that FkpA, unlike other periplasmic chaperones, increases the folded yield but decreases the folding rate of OMPs. The results indicate that FkpA behaves as a chaperone and not as a folding catalyst to influence the OMP folding trajectory. Consistent with the folding assay results, FkpA binds all three uOMPs as determined by sedimentation velocity (SV) and photo‐crosslinking experiments. We determine the binding affinity between FkpA and uOmpA171 by globally fitting SV titrations and find it to be intermediate between the known affinities of Skp and SurA for uOMP clients. Notably, complex formation steeply depends on the urea concentration, suggesting an extensive binding interface. Initial characterizations of the complex using photo‐crosslinking indicate that the binding interface spans the entire FkpA molecule. In contrast to prior findings, folding and binding experiments performed using subdomain constructs of FkpA demonstrate that the full‐length chaperone is required for full activity. Together these results support that FkpA has a distinct and direct effect on OMP folding that it achieves by utilizing an extensive chaperone‐client interface to tightly bind clients.

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Chaperone Spy Protects Outer Membrane Proteins from Folding Stress via Dynamic Complex Formation

Abstract: Outer membrane proteins (OMPs) play critical roles in bacterial pathogenicity and provide a new niche for antibiotic development. A comprehensive understanding of the OMP quality control network will strongly impact antimicrobial discovery.

Cited by 17 publications

References 58 publications

Effects of Cinnamon Essential Oil on Oxidative Damage and Outer Membrane Protein Genes of Salmonella enteritidis Cells

Effects of Cinnamon Essential Oil on Oxidative Damage and Outer Membrane Protein Genes of Salmonella enteritidis Cells

FkpA Enhances Membrane Protein Folding using an Extensive Interaction Surface

FkpA enhances membrane protein folding using an extensive interaction surface

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