Extracellular serine proteases from Stenotrophomonas maltophilia: Screening, isolation and heterologous expression in E. coli

Ribitsch, Doris; Heumann, Sonja; Karl, Wolfgang; Gerlach, Joerg C.; Leber, Regina; Birner‐Gruenberger, Ruth; Gruber, Karl; Eiteljoerg, Inge; Remler, Peter; Siegert, Petra; Lange, J.; Maurer, Karl‐Heinz; Berg, Gabriele; Guebitz, Georg M.; Schwab, Helmut

doi:10.1016/j.jbiotec.2011.09.025

Cited by 43 publications

(29 citation statements)

References 25 publications

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“…In support of our findings, StmPr1 purified from a S. maltophilia bronchial isolate hydrolyzed azocasein and N-succinyl-Ala-Ala-Pro-Phe-pNa and was sufficient to cause rounding of human fibroblasts (32). Also in line with our data, StmPr2 from the clinical isolate 19580, when purified from recombinant E. coli, exhibited caseinolytic activity (34).…”

Section: Discussionsupporting

confidence: 88%

“…StmPr1 and StmPr2 also have similar domain structures, as both proteins are predicted to exist as proenzymes and to contain a conserved catalytic triad (Asp-His-Ser) within the mature form of the protein (Fig. 2C) (32,34). Both StmPr1 and StmPr2 also have a predicted signal sequence (Fig.…”

Section: Xps T2s Causes Rounding and Detachment Of Epithelial And Fibmentioning

confidence: 95%

“…Both StmPr1 and StmPr2 also have a predicted signal sequence (Fig. 2C) (32,34), suggesting that they are substrates for a T2S system and possibly contribute to the rounding and detachment of A549 cells that is induced by strain K279a supernatant.…”

Section: Xps T2s Causes Rounding and Detachment Of Epithelial And Fibmentioning

confidence: 99%

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Type II Secretion-Dependent Degradative and Cytotoxic Activities Mediated by Stenotrophomonas maltophilia Serine Proteases StmPr1 and StmPr2

2015

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Stenotrophomonas maltophilia is an emerging opportunistic pathogen that primarily causes pneumonia and bacteremia in immunocompromised individuals. We recently reported that S. maltophilia strain K279a encodes the Xps type II secretion system and that Xps promotes rounding, actin rearrangement, detachment, and death in the human lung epithelial cell line A549. Here, we show that Xps-dependent cell rounding and detachment occur with multiple human and murine cell lines and that serine protease inhibitors block Xps-mediated rounding and detachment of A549 cells. Using genetic analysis, we determined that the serine proteases StmPr1 and StmPr2, which were confirmed to be Xps substrates, are predominantly responsible for secreted proteolytic activities exhibited by strain K279a, as well as the morphological and cytotoxic effects on A549 cells. Supernatants from strain K279a also promoted the degradation of type I collagen, fibrinogen, and fibronectin in a predominantly Xps-and protease-dependent manner, although some Xps-independent degradation of fibrinogen was observed. Finally, Xps, and predominantly StmPr1, degraded interleukin 8 (IL-8) secreted by A549 cells during coculture with strain K279a. Our findings indicate that while StmPr1 and StmPr2 are predominantly responsible for A549 cell rounding, extracellular matrix protein degradation, and IL-8 degradation, additional Xps substrates also contribute to these activities. Altogether, our data provide new insight into the virulence potential of the S. maltophilia Xps type II secretion system and its StmPr1 and StmPr2 substrates. The Gram-negative bacterium Stenotrophomonas maltophilia, prevalent in the aqueous environment, is now emerging as a prominent opportunistic and nosocomial pathogen (1, 2). S. maltophilia infects an array of host tissues and organs, including the respiratory tract, blood, bone, soft tissue, eye, urinary tract, heart, and brain. However, pneumonia is the most common infection associated with S. maltophilia, followed by bloodstream infection as the second most common (1, 2). S. maltophilia, though relatively nonvirulent for healthy individuals, can cause serious infection in immunocompromised individuals. Therefore, S. maltophilia poses the biggest threat for patients with severe burns, cystic fibrosis, and HIV, as well as those undergoing chemotherapy or immunosuppressive therapy (3). Prolonged hospitalization in intensive care units and long-term antibiotic treatment are also considered risk factors for developing pneumonia and bacteremia, for which mortality rates can range from 23 to 77% and 14 to 69%, respectively (1, 4). Community-acquired S. maltophilia infections, especially in the high-risk populations mentioned, have been reported (5), and recently, a communityacquired skin infection was reported for the first time in an immunocompetent individual (6). The multidrug-resistant nature of S. maltophilia makes treatment of infections highly difficult (7), and in recent years, an increased resistance to the preferred antibiotic trimeth...

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Section: Discussionsupporting

confidence: 88%

Section: Xps T2s Causes Rounding and Detachment Of Epithelial And Fibmentioning

confidence: 95%

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Type II Secretion-Dependent Degradative and Cytotoxic Activities Mediated by Stenotrophomonas maltophilia Serine Proteases StmPr1 and StmPr2

2015

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“…It is also an opportunistic human pathogenic bacterium (Berg et al 2005). Various beneficial applications of this Gram-negative bacterium, including biological control of fungal plant diseases, bioremediation, and the production of detergent proteases are known (Ryan et al 2009;Jin et al 2011;Ribitsch et al 2012). The present study reports for the first time the QQ and anti-biofilm activity of S. maltophilia isolated from a salt-tolerant grass species.…”

Section: Resultsmentioning

confidence: 83%

Cis-9-octadecenoic acid from the rhizospheric bacteriumStenotrophomonas maltophiliaBJ01 shows quorum quenching and anti-biofilm activities

et al. 2013

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Quorum quenching (QQ) is an effective approach for the prevention of bacterial infections involving biofilms. This study reports the QQ and anti-biofilm activities of a rhizospheric bacterium identified as Stenotrophomonas maltophilia BJ01. The QQ activity was demonstrated using Chromobacterium violaceum CV026 as a biosensor. A maximum of 95% reduction in violacein production, a quorum sensing-regulated behavior, was observed. Gas chromatography-mass spectroscopy of the extract showed that the active compound was cis-9-octadecenoic acid, which was confirmed by electronspray ionization-mass spectroscopy data. The extract also inhibited biofilm formation of Pseudomonas aeruginosa ATCC 9027 without affecting its growth. Scanning electron and atomic force microscopy showed architectural disruption of the biofilm when treated with the extract. This is the first report of the QQ and anti-biofilm activities of cis-9-octadecenoic acid isolated from any bacterium. It may have the potential to combat detrimental infections with P. aeruginosa. Further validation is required for any possible medical application.

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“…Since the pre‐peptidase C‐terminal (PPC) domain is probably related to thermostability and substrate specificity (Fang et al ., ), we hypothesized that by deleting or altering the PPC domain, we could alter the KerSMD enzyme's stability and catalytic efficiency. The catalytic efficiency of alkaline protease HP70, from Stenotrophomonas maltophilia , was found to be improved by truncation of the C‐terminal domain (Ribitsch et al ., 2010; 2012). Importantly, the KerSMD/KerSMF PPC domains and N‐propeptides have almost the same molecular size and model structure, which may improve the probability of successful exchange.…”

Section: Introductionmentioning

confidence: 97%

Enhancement of the catalytic efficiency and thermostability of Stenotrophomonas sp. keratinase KerSMD by domain exchange with KerSMF

Fang

Zhang

Liu

et al. 2015

Microbial Biotechnology

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SummaryIn this study, we enhanced the catalytic efficiency and thermostability of keratinase KerSMD by replacing its N/C‐terminal domains with those from a homologous protease, KerSMF, to degrade feather waste. Replacement of the N‐terminal domain generated a mutant protein with more than twofold increased catalytic activity towards casein. Replacement of the C‐terminal domain obviously improved keratinolytic activity and increased the kcat/Km value on a synthetic peptide, succinyl‐Ala‐Ala‐Pro‐Phe‐p‐nitroanilide, by 54.5%. Replacement of both the N‐ and C‐terminal domains generated a more stable mutant protein, with a T m value of 64.60 ± 0.65°C and a half‐life of 244.6 ± 2 min at 60°C, while deletion of the C‐terminal domain from KerSMD or KerSMF resulted in mutant proteins exhibiting high activity under mesophilic conditions. These findings indicate that the pre‐peptidase C‐terminal domain and N‐propeptide are not only important for substrate specificity, correct folding and thermostability but also support the ability of the enzyme to convert feather waste into feed additives.

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Extracellular serine proteases from Stenotrophomonas maltophilia: Screening, isolation and heterologous expression in E. coli

Cited by 43 publications

References 25 publications

Type II Secretion-Dependent Degradative and Cytotoxic Activities Mediated by Stenotrophomonas maltophilia Serine Proteases StmPr1 and StmPr2

Type II Secretion-Dependent Degradative and Cytotoxic Activities Mediated by Stenotrophomonas maltophilia Serine Proteases StmPr1 and StmPr2

Cis-9-octadecenoic acid from the rhizospheric bacteriumStenotrophomonas maltophiliaBJ01 shows quorum quenching and anti-biofilm activities

Enhancement of the catalytic efficiency and thermostability of Stenotrophomonas sp. keratinase KerSMD by domain exchange with KerSMF

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