scholarly journals Characterization of endopeptidases from the midgut of Morimus funereus (Coleoptera: Cerambycidae) larvae

2008 ◽  
Vol 60 (3) ◽  
pp. 403-409 ◽  
Author(s):  
Natasa Bozic ◽  
Biljana Dojnov ◽  
Aleksandra Milovanovic ◽  
Vera Nenadovic ◽  
Jelisaveta Ivanovic ◽  
...  
Keyword(s):  
Phenylmethylsulfonyl Fluoride ◽  
Chromogenic Substrates ◽  
Zymogram Analysis ◽  
Morimus Funereus ◽  
Specific Inhibitors

Application of specific chromogenic substrates, use of class-specific inhibitors, and zymogram analysis enabled us to identify several peptidase classes in extracts of the midgut of Morimus funereus larvae. Zymogram analysis with gelatin as a peptidase substrate and phenylmethylsulfonyl fluoride as an inhibitor showed that serine peptidases were the most abundant endopeptidases in the midgut of M. funereus larvae. By zymogram analysis with gelatin as a peptidase substrate and 1,10-phenanthroline as an inhibitor, metallopeptidases were also detected. Analyses of serine peptidases with specific chromogenic substrates revealed dominance of elastase-like peptidases in extracts of the midgut of M. funereus larvae, with less pronounced chymotrypsin- and trypsin-like activities.

Comparative characterization of digestive proteases in redhead cichlid (Vieja melanurus) and twoband cichlid (Vieja bifasciata) (Percoidei: Cichlidae)

2021 ◽  
Vol 19 (1) ◽  
Author(s):  
Carlos Alfonso Frías-Quintana ◽  
Emyr Saul Peña-Marín ◽  
Carlos David Ramírez-Custodio ◽  
Rafael Martínez-García ◽  
Luis Daniel Jiménez-Martínez ◽  
...  
Keyword(s):  
Meat Quality ◽  
Alkaline Proteases ◽  
Optimal Temperature ◽  
Digestive Capacity ◽  
Digestive Proteases ◽  
Ph Values ◽  
Specific Diet ◽  
Specific Inhibitors ◽  
Optimal Ph

ABSTRACT In the Southeast of Mexico, there are many native cichlids with commercial interest such as redhead cichlid (Vieja melanurus) and twoband cichlid (V. bifasciata), which have a great local demand and excellent meat quality. However, it is necessary to implement their culture based on nutrition studies and digestive biochemistry. This study’s objective was to characterize these two cichlids’ digestive proteases (pH, temperature, and inhibitors) through biochemistry techniques. Results showed that V. melanurus and V. bifasciata have a digestive capacity analogous to other omnivore fishes, where the optimal pH values of stomach proteases (4 and 2, respectively) and intestinal proteases (6 and 12, respectively), the optimal temperature of acid (35°C and 55°C, respectively) and alkaline proteases (45°C and 55°C, respectively) are quite similar. Both species presented high thermal and pH stabilities. Inhibition showed that V. melanurus is more sensitive to specific inhibitors for alkaline proteases than V. bifasciata. In conclusion, V. bisfasciata and V. melanurus have different digestive protease patterns. Both species can hydrolyze different protein ingredients to formulate a specific diet. Nevertheless, V. bifasciata is more resistant to the presence of inhibitors, which allow it to include vegetable proteins in its diet.

Isolation and Partial Characterization of a Novel Circulating Antithrombin

1979 ◽  
Author(s):  
Harry Messmore ◽  
Zaheer Pervez ◽  
Jawed Fareed
Keyword(s):  
Protease Inhibitor ◽  
Antithrombin Iii ◽  
Factor Xa ◽  
Serine Protease Inhibitor ◽  
Chromogenic Substrates ◽  
Inhibitor Activity ◽  
Antithrombin Activity ◽  
Life Threatening ◽  
Hydrolysis Of

We have previously reported on a novel circulating anticoagulant (Clin. Res. 22:396 A, 1974. Thromb. and Haem. 38:77, 1977 (abstr) in a 42 year old female who has had repeated episodes of life threatening hemorrhage since the age of 3. Our preliminary studies showed it to be a protein with some of the properties of a heparin activated antithrombin III. This report is on additional studies to further characterize the Inhibitor«The patient’s plasma was fractionated on Sephadex G-200, and the fraction showing immediate acting antithrombin activity was further purified on heparin-sepharose and conconavalin A-sepharose affinity columns. The patient’s inhibitor did not bind to heparin sepharose, and was thus separable from her antithrombin III. It did bind to conconavalin A, and was eluted from the conconavalin A with α - D (+) methylglucoside. It has very broad serine protease inhibitor activity, blocking the hydrolysis of synthetic chromogenic substrates by thrombin, factor Xa, plasmin and trypsin. It did not inhibit Reptilase.Immunochemical assays show it to be α1 antitrypsin. Isoelectric focusing shows it to he a variant of normal, with its isoelectric point being different from a normal control, and pure α1 antitrypsin (commercial, human).The total α1 antitrypsin level in this patient is about 50% of normal, and It has potent immediate acting antithrombin activity. It appears to be similar to a phenotype previously reported as Antithrombin Pittsburgh (Blood 51: 129, 1978).

Isolation and Partial Characterization of A Novel Circulating Antithrombin

1979 ◽  
Author(s):  
Harry Messmore ◽  
Zaheer Parvez ◽  
Jawed Fareed
Keyword(s):  
Protease Inhibitor ◽  
Antithrombin Iii ◽  
Factor Xa ◽  
Serine Protease Inhibitor ◽  
Chromogenic Substrates ◽  
Inhibitor Activity ◽  
Antithrombin Activity ◽  
Life Threatening ◽  
Hydrolysis Of

We have previously reported on a novel circulating anticoagulant (Clin. Res. 22:396 A, 1974. Thromb. and Haem. 38:77, 1977 (abstr) in a 42 year old female who has had repeated episodes of life threatening hemorrhage since the age of 3. Our preliminary studies showed it to be a protein with some of the properties of a heparin activated antithrombin III. This report is on additional studies to further characterize the inhibitor.The patient’s plasma was fractionated on Sephadex G-200, and the fraction showing immediate acting antithrombin activity was further purified on heparin-sepharose and conconavalin A-sepharose affinity columns. The patient’s inhibitor did not bind to heparin sepharose, and was thus separable from her antithrombin III. It did bind to conconavalin A, and was eluted from the conconavalin A with α - D (+) methylglucoside.It has very broad serine protease inhibitor activity, blocking the hydrolysis of synthetic chromogenic substrates by thrombin, factor Xa, plasmin and trypsin. It did not inhibit Reptilase.Immunochemical assays show it to be α1 antitrypsin. Isoelectric focusing shows it to be a variant of normal, with its isoelectric point being different from a normal control, and pure α1 antitrypsin (commercial, human).The total α1 antitrypsin level in this patient is about 50% of normal, and it has potent immediate acting antithrombin activity. It appears to be similar to a phenotype previously reported as Antithrombin Pittsburgh (Blood 51:129, 1978).

scholarly journals Peptidolytic enzymes in different larval stadium of housefly Musca domestica

2012 ◽  
Vol 51 (No. 4) ◽  
pp. 139-144 ◽  
Author(s):  
J. Blahovec ◽  
Z. Kostecka ◽  
A. Kocisova
Keyword(s):  
Enzyme Activity ◽  
Metal Ions ◽  
Larval Development ◽  
Musca Domestica ◽  
Major Part ◽  
Chelating Agent ◽  
Chromogenic Substrates ◽  
Three Stages ◽  
Specific Inhibitors

Four classes of peptidolytic enzymes were described in insects. Many authors have found predominant activity belonging to trypsin-like and chymotrypsin-like activity. By the use specific chromogenic substrates and hemoglobin we have determined enzyme activity in three stages of larval development of housefly. In contrast to above mentioned data we have found, that major part of peptidolytic activity in this insect is of aminopeptidase nature. Other observed peptidolytic activity formed only minority part. Apparently the highest activities to all examined substrates were found in first larval stadium of housefly. Inhibitory studies by class specific inhibitors and influence of metal ions and chelating agent on enzyme activity have shown, that aminopeptidase-like enzymes belong to metalloproteinase group.

Antibody production in goat milk serum after virus instillation of goat mammary gland. V. Biochemical isolation and further characterization of antibodies to various influenza and mumps viruses

1970 ◽  
Vol 16 (12) ◽  
pp. 1153-1159 ◽  
Author(s):  
Arthur E. Pasieka ◽  
L. F. Guerin ◽  
Chas. A. Mitchell
Keyword(s):  
Mammary Gland ◽  
Laboratory Animal ◽  
Goat Milk ◽  
Laboratory Animals ◽  
Lactation Period ◽  
Milk Serum ◽  
Therapeutic Uses ◽  
Biochemical Fractionation ◽  
Specific Inhibitors

This report describes the development of a method for the isolation of antibodies produced in the mammary gland and found in the milk after the instillation and propagation of various myxoviruses. Biochemical fractionation and isolation procedures have been modified and improved over our previous initial reports. The antibodies of the various influenza and mumps viruses that propagated in the gland were found to be associated with lactogammaglobulin. This report also demonstrates that the antibodies produced in the gland and thus given off in the milk are probably the same as those found in the blood if the animal were infected by conventional routes. Purification of the lactoglobulin protein fraction containing the antibody eliminated the non-specific inhibitors. These results were obtained from the various myxoviruses and mumps that propagated in the goat mammary gland.The main advantages of using the mammary gland as compared to using laboratory animals and their blood are as follows.1. Larger volumes of antibodies can be produced at one time (lactation period) against the influenza and mumps viruses for diagnostic and possibly for therapeutic uses.2. The animal (goat) does not appear to be affected whatsoever by the virus instillation and propagation techniques.3. The milk technique is therefore more humane towards laboratory animals. Invariably the laboratory animal does not have to be sacrificed.

Comparative characterization of two DEAD-box RNA helicases in superfamily II: human translation-initiation factor 4A and hepatitis C virus non-structural protein 3 (NS3) helicase

2002 ◽  
Vol 363 (1) ◽  
pp. 147-155 ◽  
Author(s):  
Mark X. DU ◽  
Robert B. JOHNSON ◽  
Xin-Lai SUN ◽  
Kirk A. STASCHKE ◽  
Joseph COLACINO ◽  
...  
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Helicase Domain ◽  
Dna Duplexes ◽  
Structural Protein ◽  
Comparative Characterization ◽  
Specific Inhibitors

Eukaryotic initiation factor 4A (eIF4A) is an ATP-dependent RNA helicase and is homologous to the non-structural protein 3 (NS3) helicase domain encoded by hepatitis C virus (HCV). Reported here is the comparative characterization of human eIF4A and HCV NS3 helicase in an effort to better understand viral and cellular helicases of superfamily II and to assist in designing specific inhibitors against HCV infections. Both eIF4A and HCV NS3 helicase domain were expressed in bacterial cells as histidine-tagged proteins and purified to homogeneity. Purified eIF4A exhibited RNA-unwinding activity and acted on RNA or RNA/DNA but not DNA duplexes. In the absence of cellular cofactors, eIF4A operated unwinding in both the 3′ to 5′ and 5′ to 3′ directions, and was able to unwind blunt-ended RNA duplex, suggesting that bidirectionality is an intrinsic property of eIF4A. In contrast, HCV NS3 helicase showed unidirectional 3′ to 5′ unwinding of RNA and RNA/DNA, as well as of DNA duplexes. With respect to NTPase activity, eIF4A hydrolysed only ATP or dATP in the presence of RNAs, whereas HCV NS3 helicase could hydrolyse all ribo- and deoxyribo-NTPs in an RNA-independent manner. In parallel, only ATP or dATP could drive the unwinding activity of eIF4A whereas HCV NS3 could function with all eight standard NTPs and dNTPs. The observed differences in their substrate specificity may prove to be useful in designing specific inhibitors targeting HCV NS3 helicase but not human eIF4A.

Characterization of three types of calcium channel in the luminal membrane of the distal nephron

2004 ◽  
Vol 82 (1) ◽  
pp. 30-37 ◽  
Author(s):  
M G Brunette ◽  
M Leclerc ◽  
D Couchourel ◽  
J Mailloux ◽  
Y Bourgeois
Keyword(s):  
Calcium Transport ◽  
Distal Tubule ◽  
Distal Nephron ◽  
High Affinity ◽  
Luminal Membrane ◽  
Kinetics Of ◽  
Substrate Concentrations ◽  
Renal Calcium Transport ◽  
Specific Inhibitors

We previously reported a dual kinetics of Ca2+ transport by the distal tubule luminal membrane of the kidney, suggesting the presence of several types of channels. To better characterize these channels, we examined the effects of specific inhibitors (i.e., diltiazem, an L-type channel; ω-conotoxin MVIIC, a P/Q-type channel; and mibefradil, a T-type channel antagonist) on 0.1 and 0.5 mM Ca2+ uptake by rabbit nephron luminal membranes. None of these inhibitors influenced Ca2+ uptake by the proximal tubule membranes. In contrast, in the absence of sodium (Na+), the three channel antagonists decreased Ca2+ transport by the distal membranes, and their action depended on the substrate concentrations: 50 µM diltiazem decreased 0.1 mM Ca2+ uptake from 0.65 ± 0.07 to 0.48 ± 0.06 pmol·µg–1·10 s–1 (P < 0.05) without influencing 0.5 mM Ca2+ transport, whereas 100 nM ω-conotoxin MVIIC decreased 0.5 mM Ca2+ uptake from 1.02 ± 0.05 to 0.90 ± 0.05 pmol·µg–1·10 s–1 (P < 0.02) and 1 µM mibefradil decreased it from 1.13 ± 0.09 to 0.94 ± 0.09 pmol·µg–1·10 s–1 (P < 0.05); the latter two inhibitors left 0.1 mM Ca2+ transport unchanged. Diltiazem decreased the Vmax of the high-affinity channels, whereas ω-conotoxin MVIIC and mibefradil influenced exclusively the Vmax of the low-affinity channels. These results not only confirm that the distal luminal membrane is the site of Ca2+ channels, but they suggest that these channels belong to the L, P/Q, and T types.Key words: renal calcium transport, calcium channels, diltiazem, mibefradil, ω-conotoxin.

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