scholarly journals Structural basis of covalent inhibitory mechanism of TMPRSS2-related serine proteases by camostat

2021 ◽  
Author(s):  
Gaohui Sun ◽  
Yaqun Sui ◽  
Yang Zhou ◽  
Junlin Ya ◽  
Cai Yuan ◽  
...  
Keyword(s):  
Protease Inhibitor ◽  
Serine Protease ◽  
Serine Proteases ◽  
Host Cells ◽  
Structural Basis ◽  
Inhibition Mechanism ◽  
Inhibitory Mechanism ◽  
Viral Pathogen ◽  
Viral Spread ◽  
Human Lung Cells

SARS-CoV-2 is the viral pathogen causing the COVID19 global pandemic. No effective treatment for COVID-19 has been established yet. TMPRSS2 is essential for viral spread and pathogenicity by facilitating the entry of SARS-CoV-2 onto host cells. The protease inhibitor camostat, an anticoagulant used in the clinic, has potential anti-inflammatory and anti-viral activities against COVID-19. However, the potential mechanisms of viral resistance and antiviral activity of camostat are unclear. Herein, we demonstrate high inhibitory potencies of camostat for a panel of serine proteases, indicating the camostat is a broad-spectrum inhibitor of serine proteases. In addition, we determined the crystal structure of camostat in complex with a serine protease (uPA), which reveals that camostat insert to the S1 pocket of uPA but was hydrolyzed by uPA, and the cleaved camostat covalently binds to the Ser195. We also generated the homology model of the structure of the TMPRSS2 serine protease domain. The model showed that camostat used the same inhibitory mechanism to inhibit the activity of TMPRSS2, and subsequently preventing SARS-CoV-2 spread. Importance section Serine proteases are a large family of enzymes critical for multiple physiological processes and proven diagnostic and therapeutic targets in several clinical indications. A serine protease transmembrane protease serine 2 (TMPRSS2) was recently found to mediate SARS-coronavirus 2 (SARS-CoV-2) entry into the host. camostat mesylate (FOY 305), a serine protease inhibitor active against TMPRSS2 and used for the treatment of oral squamous cell carcinoma and chronic pancreatitis, inhibits SARS-CoV-2 infection of human lung cells. However, the direct inhibition mechanism of camostat mesylate for TMPRSS2 is unclear. Herein, we demonstrate camostat used the same inhibitory mechanism to inhibit the activity of TMPRSS2 as uPA, and subsequently preventing SARS-CoV-2 spread.

scholarly journals Structural Basis for Dual-Inhibition Mechanism of a Non-Classical Kazal-Type Serine Protease Inhibitor from Horseshoe Crab in Complex with Subtilisin

PLoS ONE ◽  
2011 ◽  
Vol 6 (4) ◽  
pp. e18838 ◽  
Author(s):  
Rajesh T. Shenoy ◽  
Saravanan Thangamani ◽  
Adrian Velazquez-Campoy ◽  
Bow Ho ◽  
Jeak Ling Ding ◽  
...  
Keyword(s):  
Protease Inhibitor ◽  
Serine Protease ◽  
Horseshoe Crab ◽  
Serine Protease Inhibitor ◽  
Structural Basis ◽  
Inhibition Mechanism ◽  
Dual Inhibition

scholarly journals Alpha 1 Antitrypsin is an Inhibitor of the SARS-CoV-2–Priming Protease TMPRSS2

2021 ◽  
Vol 6 (1) ◽  
pp. 55-74
Author(s):  
Nurit P Azouz ◽  
Andrea Klingler ◽  
Victoria Callahan ◽  
Ivan Akhrymuk ◽  
Katarina Elez ◽  
...  
Keyword(s):  
Viral Load ◽  
Protease Inhibitor ◽  
Serine Protease ◽  
Inhibitory Activity ◽  
Serine Proteases ◽  
Serine Protease Inhibitor ◽  
Host Cells ◽  
S Protein ◽  
Relative Contribution ◽  
Alpha 1 Antitrypsin

Background: Host proteases have been suggested to be crucial for dissemination of MERS, SARS-CoV, and SARS-CoV-2 coronaviruses, but the relative contribution of membrane versus intracellular proteases remains controversial. Transmembrane serine protease 2 (TMPRSS2) is regarded as one of the main proteases implicated in the coronavirus S protein priming, an important step for binding of the S protein to the angiotensin-converting enzyme 2 (ACE2) receptor before cell entry.  Methods: We developed a cell-based assay to identify TMPRSS2 inhibitors. Inhibitory activity was established in SARS-CoV-2 viral load systems. Results: We identified the human extracellular serine protease inhibitor (serpin) alpha 1 antitrypsin (A1AT) as a novel TMPRSS2 inhibitor. Structural modeling revealed that A1AT docked to an extracellular domain of TMPRSS2 in a conformation that is suitable for catalysis, resembling similar serine protease inhibitor complexes. Inhibitory activity of A1AT was established in a SARS-CoV-2 viral load system. Notably, plasma A1AT levels were associated with COVID-19 disease severity.  Conclusions: Our data support the key role of extracellular serine proteases in SARS CoV-2 infections and indicate that treatment with serpins, particularly the FDA-approved drug A1AT, may be effective in limiting SARS-CoV-2 dissemination by affecting the surface of the host cells.

scholarly journals Carbohydrate-controlled serine protease inhibitor (serpin) production in Bifidobacterium longum subsp. longum

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
S. Duboux ◽  
M. Golliard ◽  
J. A. Muller ◽  
G. Bergonzelli ◽  
C. J. Bolten ◽  
...  
Keyword(s):  
Protease Inhibitor ◽  
Serine Protease ◽  
Serine Proteases ◽  
Intestinal Tract ◽  
Defense Mechanism ◽  
Inflammatory Diseases ◽  
Bifidobacterium Longum ◽  
Serine Protease Inhibitor ◽  
Innate Defense

AbstractThe Serine Protease Inhibitor (serpin) protein has been suggested to play a key role in the interaction of bifidobacteria with the host. By inhibiting intestinal serine proteases, it might allow bifidobacteria to reside in specific gut niches. In inflammatory diseases where serine proteases contribute to the innate defense mechanism of the host, serpin may dampen the damaging effects of inflammation. In view of the beneficial roles of this protein, it is important to understand how its production is regulated. Here we demonstrate that Bifidobacterium longum NCC 2705 serpin production is tightly regulated by carbohydrates. Galactose and fructose increase the production of this protein while glucose prevents it, suggesting the involvement of catabolite repression. We identified that di- and oligosaccharides containing galactose (GOS) and fructose (FOS) moieties, including the human milk oligosaccharide Lacto-N-tetraose (LNT), are able to activate serpin production. Moreover, we show that the carbohydrate mediated regulation is conserved within B. longum subsp. longum strains but not in other bifidobacterial taxons harboring the serpin coding gene, highlighting that the serpin regulation circuits are not only species- but also subspecies- specific. Our work demonstrates that environmental conditions can modulate expression of an important effector molecule of B. longum, having potential important implications for probiotic manufacturing and supporting the postulated role of serpin in the ability of bifidobacteria to colonize the intestinal tract.

scholarly journals Carbohydrate-controlled Serine Protease Inhibitor (Serpin) Production in Bifidobacterium Longum Subsp. Longum

2020 ◽  
Author(s):  
Stéphane Duboux ◽  
Mireille Golliard ◽  
Jeroen Muller ◽  
Gabriela Bergonzelli ◽  
Christoph Bolten ◽  
...  
Keyword(s):  
Protease Inhibitor ◽  
Serine Protease ◽  
Serine Proteases ◽  
Intestinal Tract ◽  
Defense Mechanism ◽  
Inflammatory Diseases ◽  
Bifidobacterium Longum ◽  
Serine Protease Inhibitor ◽  
Innate Defense

Abstract The Serine Protease Inhibitor (serpin) protein has been suggested to play a key role in the interaction of bifidobacteria with the host. By inhibiting intestinal serine proteases, it might allow bifidobacteria to reside in specific gut niches. In inflammatory diseases where serine proteases contribute to the innate defense mechanism of the host, serpin may dampen the damaging effects of inflammation. In view of the beneficial roles of this protein, it is important to understand how its production is regulated. Here we demonstrate that B. longum NCC 2705 serpin production is tightly regulated by carbohydrates. Galactose and fructose increase the production of this protein while glucose prevents it, suggesting the involvement of catabolite repression. We identified that di- and oligosaccharides containing galactose (GOS) and fructose (FOS) moieties, including the human milk oligosaccharide Lacto-N-tetraose (LNT), are able to activate serpin production. Moreover, we show that the carbohydrate mediated regulation is conserved within B. longum subsp. longum strains but not in other bifidobacterial taxons harboring the serpin coding gene, highlighting that the serpin regulation circuits are not only species- but also subspecies- specific. Our work demonstrates that environmental conditions can modulate expression of an important effector molecule of B. longum, having potential important implications for probiotic manufacturing and supporting the postulated role of serpin in the ability of bifidobacteria to colonize the intestinal tract.

Cartilage synthesizes the serine protease inhibitor PAI-1: Support for the involvement of serine proteases in cartilage remodeling

1991 ◽  
Vol 9 (3) ◽  
pp. 309-316 ◽  
Author(s):  
Benjamin V. Treadwell ◽  
Michele Pavia ◽  
Christine A. Towle ◽  
Vernon J. Cooley ◽  
Henry J. Mankin
Keyword(s):  
Protease Inhibitor ◽  
Serine Protease ◽  
Serine Proteases ◽  
Serine Protease Inhibitor ◽  
Pai 1

Inhibition of human kallikreins 5 and 7 by the serine protease inhibitor lympho-epithelial Kazal-type inhibitor (LEKTI)

2005 ◽  
Vol 386 (11) ◽  
pp. 1173-1184 ◽  
Author(s):  
Norman M. Schechter ◽  
Eun-Jung Choi ◽  
Zhe-Mei Wang ◽  
Yasushi Hanakawa ◽  
John R. Stanley ◽  
...  
Keyword(s):  
Protease Inhibitor ◽  
Serine Protease ◽  
Serine Proteases ◽  
Equilibrium Constants ◽  
Kinetic Studies ◽  
Serine Protease Inhibitor ◽  
Skin Development ◽  
Netherton Syndrome ◽  
Specific Association ◽  
20 Nm

Abstract LEKTI is a 120-kDa protein that plays an important role in skin development, as mutations affecting LEKTI synthesis underlie Netherton syndrome, an inherited skin disorder producing severe scaling. Its primary sequence indicates that the protein consists of 15 domains, all resembling a Kazal-type serine protease inhibitor. LEKTI and two serine proteases belonging to the human tissue kallikrein (hK) family (hK5 and hK7) are expressed in the granular layer of skin. In this study, we characterize the interaction of two recombinant LEKTI fragments containing three or four intact Kazal domains (domains 6–8 and 9–12) with recombinant rhK5, a trypsin-like protease, and recombinant rhK7, a chymotrypsin-like protease. Both fragments inhibited rhK5 similarly in binding and kinetic studies performed at pH 8.0, as well as pH 5.0, the pH of the stratum corneum where both LEKTI and proteases may function. Inhibition equilibrium constants (K i) measured either directly in concentration-dependent studies or calculated from measured association (k ass) and dissociation (k dis) rate constants were 1.2–5.5 nM at pH 8.0 and 10–20 nM at pH 5.0. At pH 8.0, k ass and k dis values were 4.7×105 M−1 s−1 and 5.5×10−4 s−1, and at pH 5.0 they were 4.0×104 M−1 s−1 and 4.3×10−4 s−1, respectively. The low K i and k dis values (t 1/2 of 20–25 min) indicate tight and specific association. Only fragment 6–9′ was a good inhibitor of rhK7, demonstrating a K i of 11 nM at pH 8.0 in a reaction that was rapidly reversible. These results show that LEKTI, at least in fragment form, is a potent inhibitor of rhK5 and that this protease may be a target of LEKTI in human skin.

In vivo contribution of serine proteases to the proteolytic activation of γENaC in aldosterone-infused rats

2012 ◽  
Vol 303 (7) ◽  
pp. F939-F943 ◽  
Author(s):  
Kohei Uchimura ◽  
Yutaka Kakizoe ◽  
Tomoaki Onoue ◽  
Manabu Hayata ◽  
Jun Morinaga ◽  
...  
Keyword(s):  
Protease Inhibitor ◽  
Serine Protease ◽  
Serine Proteases ◽  
Serine Protease Inhibitor ◽  
Proteolytic Activation ◽  
New Strategy ◽  
Salt Sensitive Hypertension ◽  
Primary Cleavage

Aldosterone plays an important role in the regulation of blood pressure by modulating the activity of the epithelial sodium channel (ENaC) that consists of α-, β-, and γ-subunits. Aldosterone induces a molecular weight shift of γENaC from 85 to 70 kDa that is necessary for the channel activation. In vitro experiments demonstrated that a dual cleavage mechanism is responsible for this shift. It has been postulated that furin executes the primary cleavage in the Golgi and that the second cleavage is provided by other serine proteases such as prostasin or plasmin at the plasma membrane. However, the in vivo contribution of serine proteases to this cleavage remains unclear. To address this issue, we administered the synthetic serine protease inhibitor camostat mesilate (CM) to aldosterone-infused rats. CM decreased the abundance of the 70-kDa form of ENaC and led to a new 75-kDa form with a concomitant increase in the urinary Na-to-K ratio. Because CM inhibits the protease activity of serine proteases such as prostasin and plasmin, but not furin, our findings strongly indicate that CM inhibited the second cleavage of γENaC and subsequently suppressed ENaC activity. The results of our current studies also suggest the possibility that the synthetic serine protease inhibitor CM might represent a new strategy for the treatment of salt-sensitive hypertension in humans.

scholarly journals Cyclic Dimeric GMP Signaling Regulates Intracellular Aggregation, Sessility, and Growth of Ehrlichia chaffeensis

2011 ◽  
Vol 79 (10) ◽  
pp. 3905-3912 ◽  
Author(s):  
Yumi Kumagai ◽  
Junji Matsuo ◽  
Zhihui Cheng ◽  
Yoshihiro Hayakawa ◽  
Yasuko Rikihisa
Keyword(s):  
Serine Protease ◽  
Serine Proteases ◽  
Matrix Protein ◽  
Response Regulator ◽  
Bacterial Biofilm ◽  
Surface Proteins ◽  
Host Cells ◽  
Ehrlichia Chaffeensis ◽  
Content Type ◽  
Bacterial Surface

ABSTRACTCyclic dimeric GMP (c-di-GMP), a bacterial second messenger, is known to regulate bacterial biofilm and sessility. Replication of an obligatory intracellular pathogen,Ehrlichia chaffeensis, is characterized by formation of bacterial aggregates called morulae inside membrane-bound inclusions. WhenE. chaffeensismatures into an infectious form, morulae become loose to allow bacteria to exit from host cells to infect adjacent cells.E. chaffeensisexpresses a sensor kinase, PleC, and a cognate response regulator, PleD, which can produce c-di-GMP. A hydrophobic c-di-GMP antagonist, 2′-O-di(tert-butyldimethysilyl)-c-di-GMP (CDGA) inhibitsE. chaffeensisinternalization into host cells by facilitating degradation of some bacterial surface proteins via endogenous serine proteases. In the present study, we found that PleC and PleD were upregulated synchronously during exponential growth of bacteria, concomitant with increased morula size. While CDGA did not affect host cells, when infected cells were treated with CDGA, bacterial proliferation was inhibited, morulae became less compact, and the intracellular movement of bacteria was enhanced. Concurrently, CDGA treatment facilitated the extracellular release of bacteria with lower infectivity than those spontaneously released from sham-treated cells. Addition of CDGA to isolated inclusions induced dispersion of the morulae, degradation of an inclusion matrix protein TRP120, and bacterial intrainclusion movement, all of which were blocked by a serine protease inhibitor. These results suggest that c-di-GMP signaling regulates aggregation and sessility ofE. chaffeensiswithin the inclusion through stabilization of matrix proteins by preventing the serine protease activity, which is associated with bacterial intracellular proliferation and maturation.

Sign in / Sign up

Export Citation Format

Share Document