Dataset Information

Functional Proteomic Profiling of Secreted Serine Proteases in Health and Inflammatory Bowel Disease.

ABSTRACT: While proteases are essential in gastrointestinal physiology, accumulating evidence indicates that dysregulated proteolysis plays a pivotal role in the pathophysiology of inflammatory bowel disease (IBD). Nonetheless, the identity of overactive proteases released by human colonic mucosa remains largely unknown. Studies of protease abundance have primarily investigated expression profiles, not taking into account their enzymatic activity. Herein we have used serine protease-targeted activity-based probes (ABPs) coupled with mass spectral analysis to identify active forms of proteases secreted by the colonic mucosa of healthy controls and IBD patients. Profiling of (Pro-Lys)-ABP bound proteases revealed that most of hyperactive proteases from IBD secretome are clustered at 28-kDa. We identified seven active proteases: the serine proteases cathepsin G, plasma kallikrein, plasmin, tryptase, chymotrypsin-like elastase 3?A, and thrombin and the aminopeptidase B. Only cathepsin G and thrombin were overactive in supernatants from IBD patient tissues compared to healthy controls. Gene expression analysis highlighted the transcription of genes encoding these proteases into intestinal mucosae. The functional ABP-targeted proteomic approach that we have used to identify active proteases in human colonic samples bears directly on the understanding of the role these enzymes may play in the pathophysiology of IBD.

SUBMITTER: Denadai-Souza A

PROVIDER: S-EPMC5959920 | biostudies-literature | 2018 May

REPOSITORIES: biostudies-literature

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Functional Proteomic Profiling of Secreted Serine Proteases in Health and Inflammatory Bowel Disease.

Denadai-Souza Alexandre A Bonnart Chrystelle C Tapias Núria Solà NS Marcellin Marlène M Gilmore Brendan B Alric Laurent L Bonnet Delphine D Burlet-Schiltz Odile O Hollenberg Morley D MD Vergnolle Nathalie N Deraison Céline C

Scientific reports 20180518 1

While proteases are essential in gastrointestinal physiology, accumulating evidence indicates that dysregulated proteolysis plays a pivotal role in the pathophysiology of inflammatory bowel disease (IBD). Nonetheless, the identity of overactive proteases released by human colonic mucosa remains largely unknown. Studies of protease abundance have primarily investigated expression profiles, not taking into account their enzymatic activity. Herein we have used serine protease-targeted activity-base ...[more]

PMID: 29777136

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Project description:In eukaryotes, the serpins constitute a wide family of protease inhibitors regulating many physiological pathways. Many reports stressed the key role of serpins in several human physiopathologies including mainly the inflammatory bowel diseases. In this context, eukaryotic serpins were largely studied and their use to limit inflammation was reported. In comparison to that, bacterial serpins and mainly those from human gut microbiota remain poorly studied.The two genes encoding for putative serpins from the human gut bacterium Eubacterium sireaum, display low sequence identities. These genes were overexpressed and the encoded proteins, named Siropins, were purified. Activity studies demonstrated that both purified proteins inhibited serine proteases but surprisingly they preferentially inhibited two human serine proteases (Human Neutrophil Elastase and Proteinase3). The biochemical characterization of these Siropins revealed that Siropin 1 was the most active and stable at low pH values while Siropin 2 was more thermoactive and thermostable. Kinetic analysis allowed the determination of the stoichiometry of inhibition (SI) which was around 1 and of the association rate constants of 7.7 × 104 for the Human Neutrophil Elastase and 2.6 × 105 for the Proteinase3. Moreover, both Siropins displayed the ability to inhibit proteases usually present in fecal waters. Altogether our data indicate the high efficiency of Siropins and their probable involvement in the control of the overall intestine protease activity.Here we report the purification and the biochemical characterization of two novel serpins originated from Eubacterium sireaum, a human gastro-intestinal tract commensal bacteria. These proteins that we called Siropins, efficiently inhibited two human proteases reported to be associated with inflammatory bowel diseases. The determination of the biochemical properties of these enzymes revealed different temperature and pH behaviours that may reflect adaptation of this human commensal bacterium to different ecological environments. To the best of our knowledge, it is the first bacterial serpins showing an attractive inhibition of fecal proteases recovered from a mice group with chemically induced inflammation. Altogether our data highlight the interesting potential of Siropins, and serpins from the human gut microbiota in general, to be used as new alternative to face inflammatory diseases.

Project description:This review provides a summary of the global epidemiology of inflammatory bowel diseases (IBD). It is now clear that IBD is increasing worldwide and has become a global emergence disease. IBD, which includes Crohn's disease (CD) and ulcerative colitis (UC), has been considered a problem in industrial-urbanized societies and attributed largely to a Westernized lifestyle and other associated environmental factors. Its incidence and prevalence in developing countries is steadily rising and has been attributed to the rapid modernization and Westernization of the population. There is a need to reconcile the most appropriate treatment for these patient populations from the perspectives of both disease presentation and cost. In the West, biological agents are the fastest-growing segment of the prescription drug market. These agents cost thousands of dollars per patient per year. The healthcare systems, and certainly the patients, in developing countries will struggle to afford such expensive treatments. The need for biological therapy will inevitably increase dramatically, and the pharmaceutical industry, healthcare providers, patient advocate groups, governments and non-governmental organizations should come to a consensus on how to handle this problem. The evidence that IBD is now affecting a much younger population presents an additional concern. Meta-analyses conducted in patients acquiring IBD at a young age also reveals a trend for their increased risk of developing colorectal cancer (CRC), since the cumulative incidence rates of CRC in IBD-patients diagnosed in childhood are higher than those observed in adults. In addition, IBD-associated CRC has a worse prognosis than sporadic CRC, even when the stage at diagnosis is taken into account. This is consistent with additional evidence that IBD negatively impacts CRC survival. A continuing increase in IBD incidence worldwide associated with childhood-onset of IBD coupled with the diseases' longevity and an increase in oncologic transformation suggest a rising disease burden, morbidity, and healthcare costs. IBD and its associated neoplastic transformation appear inevitable, which may significantly impact pediatric gastroenterology and adult CRC care. Due to an infrastructure gap in terms of access to care between developed vs. developing nations and the uneven representation of IBD across socioeconomic strata, a plan is needed in the developing world regarding how to address this emerging problem.

Dataset Information

Functional Proteomic Profiling of Secreted Serine Proteases in Health and Inflammatory Bowel Disease.

Publications

Functional Proteomic Profiling of Secreted Serine Proteases in Health and Inflammatory Bowel Disease.

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