scholarly journals Activity of serine proteases from Fasciola hepatica eggs in relation to pH and temperature

2020 ◽  
Vol 23 (3) ◽  
pp. 350-358
Author(s):  
L. Kianifard ◽  
M. Yakhchali ◽  
M. Imani
Keyword(s):  
Serine Protease ◽  
Protease Inhibitors ◽  
Proteolytic Activity ◽  
Serine Proteases ◽  
Fasciola Hepatica ◽  
Ethylenediaminetetraacetic Acid ◽  
Optimum Temperature ◽  
Egg Hatching ◽  
Protein Levels ◽  
Ph Values

This study was conducted to analyse the serine protease of Fasciola hepatica eggs by specific substrates and inhibitors, and investigation of the effects of pH and temperature on proteases’ activity and stability. Adult worms were isolated from infected livers. After homogenisation, their protein levels were measured with the Bradford method. Total proteolytic activity of the Fasciola hepatica extract was evaluated with azocasein substrate at pH values from 2 to 12. N-benzoyl–arginine–p-nitroanilide (BApNA) trypsin and N-succinyl-alanine-alanine-prolin-phenylalanine-p-nitroanilide (SAAPFpNA) chymotrypsin substrates were used to measure specific protease activities. The effect of protease inhibitors phenylmethane sulfonyl fluoride (PMSF), pepsin, and ethylenediaminetetraacetic acid (EDTA) on these enzymes was evaluated. Estimation of optimum temperature and pH was performed in the temperature range of 10–90 °C and pH values from 2–12. The optimum pH activities for trypsin and chymotrypsin were at alkaline pH and for total proteolytic activity at acidic pH. The results using protease inhibitors showed that the eggs had serine protease activity. The optimum temperature activity of trypsin and chymotrypsin was 50 °C. These proteases were stable up to 40 °C. Due to the importance of pH and temperature in life cycle of Fasciola hepatica, these findings can be used for induction of some modifications in pH and preventing the activity of the enzyme for decrement of the efficacy of embryonic development and egg hatching of this zoonotic parasite.

scholarly journals The Effects of pH and Temperature on Cysteine Protease (Cathepsin B) Activity in Miracidia and Eggs of Fasciola hepatica

2020 ◽  
Author(s):  
Leila KIANIFARD ◽  
Mohammad YAKHCHALI ◽  
Mehdi IMANI
Keyword(s):  
Cysteine Protease ◽  
Cathepsin B ◽  
Fasciola Hepatica ◽  
Distilled Water ◽  
Acidic Ph ◽  
Optimum Temperature ◽  
Egg Hatching ◽  
Ph Values ◽  
Zoonotic Parasite ◽  
Effects Of Ph

Abstract Background: Fascioliasis is a worldwide zoonotic disease caused by the trematodes Fasciola hepatica in humans and animals. Proteases are essential for the survival of parasites and have important activities such as penetration, tissue migration, and egg hatching. This study was conducted to analyze cysteine protease of the miracidia and eggs of F. hepatica, and to assess the effects of pH and temperature on the proteases activity and stability. Methods: Adults F. hepatica were isolated from infected livers and were morphologically identified in 2018. Eggs collected from the adults and incubated in distilled water at 28 °C for 16 d to produce miracidia. The extract was collected from miracidia and eggs. A substrate for cathepsin B (Z-Arg-Arg-Pna) was used to assess the enzyme activity at different (2-12) pH levels. After homogenization, protein level was measured with Bradford method. Estimation of optimum temperature and pH was performed in the temperature range of 10-90 ° C and pH values from 2-12. Results: The highest activity of the miracidia and eggs enzyme extracts for Z-ArgArg-pNA was at pH 4. The miracidia extract was most stable at neutral pH and the eggs extract was most stable in acidic pH. The optimum temperature activity for both stages was 40 °C. These proteases were stable up to 40 °C. Conclusion: Upon the importance of pH and temperature in the life cycle of F. hepatica, the current findings can be used for induction of some modifications in pH and preventing the activity of the enzymes for decrement of the efficacy of miracidia penetration into the intermediate snails and egg hatching of this zoonotic parasite.

scholarly journals Hindrance of the Proteolytic Activity of Neutrophil-Derived Serine Proteases by Serine Protease Inhibitors as a Management of Cardiovascular Diseases and Chronic Inflammation

2021 ◽  
Vol 9 ◽  
Author(s):  
Timo Burster ◽  
Zhadyra Mustafa ◽  
Dinara Myrzakhmetova ◽  
Anuar Zhanapiya ◽  
Michal Zimecki
Keyword(s):  
Cardiovascular Diseases ◽  
Serine Protease ◽  
Chronic Inflammation ◽  
Protease Inhibitors ◽  
Proteolytic Activity ◽  
Serine Proteases ◽  
Logical Consequence ◽  
Immune Defense ◽  
Serine Protease Inhibitors ◽  
Surrounding Environment

During inflammation neutrophils become activated and segregate neutrophil serine proteases (NSPs) to the surrounding environment in order to support a natural immune defense. However, an excess of proteolytic activity of NSPs can cause many complications, such as cardiovascular diseases and chronic inflammatory disorders, which will be elucidated on a biochemical and immunological level. The application of selective serine protease inhibitors is the logical consequence in the management of the indicated comorbidities and will be summarized in this briefing.

scholarly journals Serine proteases and serine protease inhibitors in testicular physiology: the plasminogen activation system

Reproduction ◽  
2007 ◽  
Vol 134 (6) ◽  
pp. 721-729 ◽  
Author(s):  
Brigitte Le Magueresse-Battistoni
Keyword(s):  
Serine Protease ◽  
Protease Inhibitors ◽  
Expression Pattern ◽  
Serine Proteases ◽  
Current Knowledge ◽  
Adult Life ◽  
Plasminogen Activation ◽  
Serine Protease Inhibitors ◽  
Plasminogen Activation System ◽  
Activation System

The testis is an organ in which a series of radical remodeling events occurs during development and in adult life. These events likely rely on a sophisticated network of proteases and complementary inhibitors, including the plasminogen activation system. This review summarizes our current knowledge on the testicular occurrence and expression pattern of members of the plasminogen activation system. The various predicted functions for these molecules in the establishment and maintenance of the testicular architecture and in the process of spermatogenesis are presented.

scholarly journals A Versatile and Robust Serine Protease Inhibitor Scaffold from Actinia tenebrosa

Marine Drugs ◽  
2019 ◽  
Vol 17 (12) ◽  
pp. 701 ◽  
Author(s):  
Xingchen Chen ◽  
Darren Leahy ◽  
Jessica Van Haeften ◽  
Perry Hartfield ◽  
Peter J. Prentis ◽  
...  
Keyword(s):  
Serine Protease ◽  
Protease Inhibitors ◽  
Serine Proteases ◽  
Inhibitor Design ◽  
Consensus Analysis ◽  
Fine Grained ◽  
Human Sequence ◽  
Physiological Processes ◽  
Definition Of ◽  
Dynamics Simulations

Serine proteases play pivotal roles in normal physiology and a spectrum of patho-physiological processes. Accordingly, there is considerable interest in the discovery and design of potent serine protease inhibitors for therapeutic applications. This led to concerted efforts to discover versatile and robust molecular scaffolds for inhibitor design. This investigation is a bioprospecting study that aims to isolate and identify protease inhibitors from the cnidarian Actinia tenebrosa. The study isolated two Kunitz-type protease inhibitors with very similar sequences but quite divergent inhibitory potencies when assayed against bovine trypsin, chymostrypsin, and a selection of human sequence-related peptidases. Homology modeling and molecular dynamics simulations of these inhibitors in complex with their targets were carried out and, collectively, these methodologies enabled the definition of a versatile scaffold for inhibitor design. Thermal denaturation studies showed that the inhibitors were remarkably robust. To gain a fine-grained map of the residues responsible for this stability, we conducted in silico alanine scanning and quantified individual residue contributions to the inhibitor’s stability. Sequences of these inhibitors were then used to search for Kunitz homologs in an A. tenebrosa transcriptome library, resulting in the discovery of a further 14 related sequences. Consensus analysis of these variants identified a rich molecular diversity of Kunitz domains and expanded the palette of potential residue substitutions for rational inhibitor design using this domain.

scholarly journals Insights into the regulation of the matriptase-prostasin proteolytic system

2020 ◽  
Vol 477 (22) ◽  
pp. 4349-4365
Author(s):  
Lasse Holt-Danborg ◽  
Signe Skovbjerg ◽  
Kristian W. Goderum ◽  
Annika W. Nonboe ◽  
Evelina Stankevic ◽  
...  
Keyword(s):  
Protease Inhibitors ◽  
Proteolytic Activity ◽  
Strong Evidence ◽  
Serine Proteases ◽  
Proteolytic System ◽  
Proteolytic Pathway ◽  
Epithelial Development ◽  
Dual Action ◽  
The Common

The membrane-associated prostasin and matriptase belonging to the S1A subfamily of serine proteases, are critical for epithelial development and maintenance. The two proteases are involved in the activation of each other and are both regulated by the protease inhibitors, HAI-1 and HAI-2. The S1A subfamily of serine proteases are generally produced as inactive zymogens requiring a cleavage event to obtain activity. However, contrary to the common case, the zymogen form of matriptase exhibits proteolytic activity, which can be inhibited by HAI-1 and HAI-2, as for the activated counterpart. We provide strong evidence that also prostasin exhibits proteolytic activity in its zymogen form. Furthermore, we show that the activity of zymogen prostasin can be inhibited by HAI-1 and HAI-2. We report that zymogen prostasin is capable of activating zymogen matriptase, but unable to activate its own zymogen form. We propose the existence of an unusual enzyme–enzyme relationship consisting of proteolytically active zymogen forms of both matriptase and prostasin, kept under control by HAI-1 and HAI-2, and located at the pinnacle of an important proteolytic pathway in epithelia. Perturbed balance in this proteolytic system is likely to cause rapid and efficient activation of matriptase by the dual action of zymogen matriptase and zymogen prostasin. Previous studies suggest that the zymogen form of matriptase performs the normal proteolytic functions of the protease, whereas excess matriptase activation likely causes carcinogenesis. HAI-1 and HAI-2 are thus important for the prevention of matriptase activation whether catalysed by zymogen/activated prostasin (this study) or zymogen/activated matriptase (previous studies).

scholarly journals Therapeutic targeting of coronavirus spike glycoprotein priming

2020 ◽  
Author(s):  
Maurizio Pellecchia ◽  
Elisa Barile ◽  
Carlo Baggio ◽  
Luca Gambini ◽  
Sergey A. Shiryaev ◽  
...  
Keyword(s):  
Serine Protease ◽  
Protease Inhibitors ◽  
Serine Proteases ◽  
Viral Proteins ◽  
Exact Nature ◽  
Cleavage Sites ◽  
Serine Protease Inhibitors ◽  
Spike Glycoprotein ◽  
Highly Pathogenic

Abstract Processing of certain viral proteins and bacterial toxins by host serine proteases is a frequent and critical step in virulence. The coronavirus spike glycoprotein contains three (S1, S2, and S2’) cleavage sites that are processed by human host proteases. The exact nature of these cleavage sites, and their respective processing proteases, can determine whether the virus can cross species, and the level of pathogenicity. Recent comparisons of the genomes of the highly pathogenic SARS-CoV2 and MERS-CoV, with less pathogenic strains (e.g., Bat-RaTG13, the bat homologue of SARS-CoV2) identified possible mutations in the receptor binding domain and in the S1 and S2’ cleavage sites of their spike glycoprotein. However there remains some confusion on the relative roles of the possible serine-proteases involved for priming. Using anthrax toxin as a model system, we show that in vivo inhibition of priming by pan-active serine protease inhibitors can be effective at suppressing virulence. Hence, our studies should encourage further efforts in developing either pan-serine protease inhibitors or inhibitor cocktails to target SARS-CoV2 and potentially ward off future pandemics that could develop because of the additional mutations in the S-protein priming sequence in coronaviruses.

Alpha-1 Antitrypsin Is a Potent Inhibitor of Cytokine-Induced Hematopoietic Stem Cell Mobilization.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 1185-1185
Author(s):  
Melissa van Pel ◽  
Ronald van Os ◽  
Gerjo A. Velders ◽  
Henny Hagoort ◽  
Ivan J. Lindley ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Serine Protease ◽  
Protease Inhibitors ◽  
Serine Proteases ◽  
Low Dose ◽  
Serine Protease Inhibitors ◽  
Hematopoietic Stem ◽  
Substrate Conversion

Abstract Previously, we have shown that IL-8 and G-CSF-induced hematopoietic stem cell (HSC) mobilization is inhibited in mice that underwent low dose (0.5 Gy) total body irradiation (TBI), whereas the number of progenitor cells in the bone marrow remained similar in all groups. The mechanism underlying this inhibition remains unknown. Since the release of granular proteases by neutrophils is well known to play a role in HSC mobilization, we also considered a possible role for serine protease inhibitors in the induction of HSC mobilization. Serine proteases, such as elastase and cathepsin G, are irreversibly inhibited by serine protease inhibitors including alpha-1 antitrypsin (alpha-1 AT) and alpha2-macroglobulin. In-vitro tests revealed that addition of bone marrow extracellular extracts, that were obtained from murine femurs 24 hours following low dose (0.5 Gy) TBI, inhibited the activity of exogenous elastase in a chromogenic substrate conversion assay up to 78.1 % compared to extracts obtained from sham irradiated controls (p<0.05). Since elastase inhibition by alpha2-macroglobulin cannot be detected in a chromogenic substrate conversion assay, alpha-1 AT was considered as the primary candidate serine protease inhibitor to inhibit elastase activity in our in-vitro system. Quantitative PCR of total bone marrow cells revealed that alpha-1 AT mRNA was 20-fold increased relative to the housekeeping gene ß-actin and 7-fold relative to the housekeeping genes HPRT and GAPDH at 24 hours following low dose (0.5 Gy) TBI. In addition, Western blot analysis indicated that alpha-1 AT protein concentrations were significantly (p<0.01) increased in bone marrow extracellular extracts derived from low dose (0.5 Gy) irradiated mice, compared to extracts obtained from sham-irradiated controls (5.1 ± 0.6 scanning units [SU] vs. 3.9 ± 0.7 SU for 0.5 Gy;n=8 vs. 0 Gy; n=6 respectively). To further substantiate a possible in-vivo role of alpha-1 AT in the inhibition of HSC mobilization, we administered alpha-1 AT (300 μg/mouse i.p.) at 2 hours and at 5 minutes prior to IL-8 injection (30 μg/mouse i.p.). Administration of alpha-1 AT prior to IL-8 injection completely (p<0.05) inhibited IL-8-induced HSC mobilization (472.9 ± 289.5 CFU-GM per ml blood for IL-8; n=5 vs. 44.8 ± 35.5 CFU-GM per ml blood for alpha-1 AT/IL-8; n =11). These results indicate that 1) alpha-1 AT is a potent inhibitor of IL-8-induced HSC mobilization and 2) in-vivo induced alpha-1 AT contributes to the inhibition of HSC mobilization after low-dose (0.5 Gy) TBI. We hypothesize that a critical balance between serine proteases and serine protease inhibitors plays an important role in cytokine-induced HSC mobilization.

Regulation of kallikrein-related peptidases in the skin – from physiology to diseases to therapeutic options

2013 ◽  
Vol 110 (09) ◽  
pp. 442-449 ◽  
Author(s):  
Jan Fischer ◽  
Ulf Meyer-Hoffert
Keyword(s):  
Serine Protease ◽  
Protease Inhibitors ◽  
Therapeutic Intervention ◽  
Serine Proteases ◽  
Current Knowledge ◽  
Serine Protease Inhibitors ◽  
Specific Expression ◽  
Physiological Processes ◽  
Promising Target ◽  
Protease Activation

SummaryKallikrein-related peptidases (KLKs) constitute a family of 15 highly conserved serine proteases, which show a tissue-specific expression profile. This made them valuable tumour expression markers. It became evident that KLKs are involved in many physiological processes like semen liquefaction and skin desquamation. More recently, we have learnt that they are involved in many pathophysiological conditions and diseases making them promising target of therapeutic intervention. Therefore, regulation of KLKs raised the interest of numerous reports. Herein, we summarise the current knowledge on KLKs regulation with an emphasis on skin-relevant KLKs regulation processes. Regulation of KLKs takes place on the level of transcription, on protease activation and on protease inactivation. A variety of protease inhibitors has been described to interact with KLKs including the irreversible serine protease inhibitors (SERPINs) and the reversible serine protease inhibitors of Kazal-type (SPINKs). In an attempt to integrate current knowledge, we propose that KLK regulation has credentials as targets for therapeutic intervention.

Bacterial Protease Enzyme Preparations for the Production of Non-Bitter Protein Hydrolysates

2020 ◽  
Vol 36 (4) ◽  
pp. 42-48
Author(s):  
A.S. Sereda ◽  
I.A. Velikoretskaya ◽  
D.T. Mineeva ◽  
N.V. Tsurikova ◽  
E.A. Rubtsova
Keyword(s):  
Bacillus Subtilis ◽  
Serine Protease ◽  
Proteolytic Activity ◽  
Serine Proteases ◽  
Basic Research ◽  
Culture Liquid ◽  
Enzyme Preparations ◽  
Protease Enzyme ◽  
Almost All ◽  
Casein Hydrolysates

Casein hydrolysates were obtained using enzyme preparations (EPs) of bacterial proteases, Alcalase, Protamex, Neutrase, and Protosubtilin. The dependence of the hydrolysate bitterness on the decrease of the EP proteolytic activity in the presence of PMSF, an inhibitor of serine proteases, was observed. It was found that the use of Neutrase provided non-bitter hydrolysates, while the proteolytic activity of Neutrase decreased by 44% in the presence of PMSF. A domestic EP of Protosubtilin lost 59% activity in the presence of PMSF and formed bitter hydrolysates. Casein hydrolysates obtained using Alcalase were characterized by the most severe bitterness, probably due to the fact that almost all proteolytic activity in this preparation is represented by a serine protease. The Bacillus subtilis 359 strain with a ratio of neutral and serine protease activities of 52 to 48% was selected from the VNIIPBT Collection of Bacterial Producers of Proteases. The concentrated preparation was obtained from the culture liquid of B. subtilis 359 strain by spray-drying. The preparation obtained hydrolyzed casein similarly to Neutrase and ensured the absence of bitterness in hydrolysates. neutral protease, serine protease, bitterness of hydrolysates, Bacillus subtilis This work was financially supported by a Grant of the Basic Research Program of the Russian Academy of Sciences for 2019-2021 (Project no. 0529-2019-0066).

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