Kinetic mechanism of protease inhibition by α1-antitrypsin

Kang, Un Beom; Baek, Je Hyun; Ryu, Seung Hyun; Kim, Joon; Yu, Myeong Hee; Lee, Cheolju

doi:10.1016/j.bbrc.2004.08.105

Cited by 14 publications

(17 citation statements)

References 25 publications

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“…3b, c). SI values of Siropins with HNE are lower than that of serpin from Tannerella forsythia (SI = 3.4) and similar to those of serpins from B. longum , T. tengcondensis and of the human α-1-antitrypsin [13, 14, 24]. Concerning the PR3, the SIs of Siropins were similar to that of the human α-1-antitrypsin [25].…”

Section: Resultsmentioning

confidence: 90%

Siropins, novel serine protease inhibitors from gut microbiota acting on human proteases involved in inflammatory bowel diseases

et al. 2016

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BackgroundIn 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.ResultsThe 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.ConclusionsHere 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.Electronic supplementary materialThe online version of this article (doi:10.1186/s12934-016-0596-2) contains supplementary material, which is available to authorized users.

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Section: Resultsmentioning

confidence: 90%

Siropins, novel serine protease inhibitors from gut microbiota acting on human proteases involved in inflammatory bowel diseases

et al. 2016

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“…In the current study, the concentration increase of alpha1-antitrypsin presents in the elevated appearances of the truncated forms or fragments of the protein in gastritis sera. Since alpha1-antitrypsin is a single-chain 52-kDa protein, the protein spots detected at around 40 kDa are probably the proteolytically modified molecules having lost antitryptic activity [32][33][34]. Other fragments appear in even lower MW regions (25-30 kDa) spread out in all different pI areas.…”

Section: Discussionmentioning

confidence: 97%

Serological Proteomics of Gastritis: Degradation of Apolipoprotein A-I and Alpha1-Antitrypsin Is a Common Response to Inflammation Irrespective of Helicobacter pylori Infection

Huai-yi

Wong

et al. 2008

Dig Dis Sci

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Proteomic technology was employed to analyze serum samples from healthy subjects (10 cases) and gastritis patients with negative and positive Helicobacter pylori (Hp) infection (15 cases each). The serum proteins were separated by two-dimensional (2-D) gel electrophoresis and analyzed by a computer-aided program. The altered proteins in expression were then identified by mass spectrometry and validated by Western blotting. Compared to those in normal control, proteins in at least six areas of 2-D gels were found to significantly increase their expression levels in both Hp-negative and Hp-positive serum samples. These proteins were identified by mass peptide fingerprinting and confirmed by Western blotting to be the truncated or cleaved protein fragments of apolipoprotein A-I and alpha-1 antitrypsin, two well-known acute-phase proteins. We conclude that the degradation or metabolization of acute-phase proteins, apolipoprotein A-I, and alpha1-antitrypsin, is a common response to gastric inflammation irrespective of Hp infection.

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“…In this study, we developed a small peptide with the constrained reactive site loop (RSL) structure of bioengineered serpin PDX as a modulator of furin activity. It was already demonstrated that RSL of all serpins are prime interactive domains responsible for their protease inhibitory function [20][21][22][23]. In this paper, we describe a linear and a corresponding disulfide-bridged cyclic peptide variant based on RSL structure of a1-PDX and demonstrated that both inhibit furin activity but with differential manner.…”

Section: Introductionmentioning

confidence: 89%

Modulating furin activity with designed mini‐PDX peptides: Synthesis and in vitro kinetic evaluation

Basak¹,

Lotfipour²

2005

FEBS Letters

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A peptide was designed from reactive site loop structure of a1 Antitrypsin Portland known as a1 PDX as a novel mini-PDX inhibitor of furin. The sequence was derived from (367-394) that contains the crucial furin cleavage motif RIPR 382 . A P3 mutant replacing Ile 380 by Leu was prepared as a first model peptide. A Cys residue was inserted at each terminal of the peptide for purpose of cyclisation which was accomplished by air or iodine-induced oxidation. This mini-PDX peptide both cyclic and acyclic form inhibited in vitro furin activity (IC 50 in nM) when measured against either substrates Boc-RVRRßMCA or QVEGF-C [Abz-QVHSIIRRßSLP-Y(NO 2 )-A-CONH 2 , Abz = 2-amino benzoic acid and Y(NO 2 ) = 3-nitro tyrosine], latter being derived from vascular endothelial growth factor-C (VEGF-C) processing site. The geometrically constrained structure mimicking PDX reactive loop is crucial for enzyme inhibition. Our study further revealed that both mini-PDX peptides inactivate furin in a slow tight binding manner, with disulfidebridged cyclic form being slightly more potent. Unlike PDX, these peptides inhibit furin via a different mechanistic pathway. The study provides an alternate strategy for development of efficient peptide-based inhibitors of Proprotein Convertases including furin.

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Kinetic mechanism of protease inhibition by α1-antitrypsin

Cited by 14 publications

References 25 publications

Siropins, novel serine protease inhibitors from gut microbiota acting on human proteases involved in inflammatory bowel diseases

Siropins, novel serine protease inhibitors from gut microbiota acting on human proteases involved in inflammatory bowel diseases

Serological Proteomics of Gastritis: Degradation of Apolipoprotein A-I and Alpha1-Antitrypsin Is a Common Response to Inflammation Irrespective of Helicobacter pylori Infection

Modulating furin activity with designed mini‐PDX peptides: Synthesis and in vitro kinetic evaluation

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