scholarly journals Studies on the inhibition of ferrochelatase by N-alkylated dicarboxylic porphyrins. Steric factors involved and evidence that the inhibition is reversible

1985 ◽  
Vol 226 (2) ◽  
pp. 537-544 ◽  
Author(s):  
F De Matteis ◽  
A H Gibbs ◽  
C Harvey
Keyword(s):  
Enzyme Inhibition ◽  
Active Site ◽  
Inhibitory Activity ◽  
Alkyl Group ◽  
Structural Requirements ◽  
Prolonged Incubation ◽  
Enzyme Active Site ◽  
Steric Factors ◽  
Haem Oxygenase ◽  
Pyrrole Nitrogen

The structural requirements for the inhibition of ferrochelatase by N-alkylated porphyrins were investigated and experiments carried out to explore the mechanism of enzyme inhibition. Three dicarboxylic porphyrins, all substrates of the enzyme, are strongly inhibitory when N-alkylated; in contrast, uroporphyrin and coproporphyrin (which are not substrates) do not inhibit after N-alkylation. Free carboxylic acid functions are required for inhibition, as the methyl ester derivatives are not themselves inhibitory. Porphyrins bearing the alkyl group on the pyrrole nitrogen of rings C and D are less effective inhibitors, particularly when zinc is chelated in the centre of the tetrapyrrole or the N-alkyl group is relatively large in size. The substituents at the 2- and 4-positions of the porphyrin system may also affect the inhibitory activity, particularly for the isomers with ring C and D alkylated. The zinc chelates of several N-alkylprotoporphyrins are inhibitory towards haem oxygenase, another haem-binding enzyme, and also in this case increasing the size of the alkyl group decreased the inhibitory activity, particularly for isomers with ring C or D alkylated. The inhibition could be reversed by prolonged incubation with excess porphyrin substrate, but dealkylation of the N-alkylporphyrin during enzyme inhibition could not be demonstrated. It is concluded (a) that N-alkylated dicarboxylic porphyrins compete reversibly with the porphyrin substrate for the enzyme active site and (b) that the structural and steric factors discussed above affect the inhibitory activity by modifying the affinity of the N-alkylporphyrin inhibitor for the enzyme.

scholarly journals The Physical Adsorption of Gelatinized Starch with Tannic Acid Decreases the Inhibitory Activity of the Polyphenol against α-Amylase

Foods ◽  
2021 ◽  
Vol 10 (6) ◽  
pp. 1233
Author(s):  
Yueyi Wang ◽  
Shuangshuang Li ◽  
Fangting Bai ◽  
Junwei Cao ◽  
Lijun Sun
Keyword(s):  
Enzyme Inhibition ◽  
Tannic Acid ◽  
Inhibitory Activity ◽  
Adsorption Property ◽  
Physical Adsorption ◽  
Enzyme Active Site ◽  
Opposite Trend ◽  
Gelatinized Starch ◽  
The Difference

The effects of mixing orders of tannic acid (TA), starch, and α-amylase on the enzyme inhibition of TA were studied, including mixing TA with α-amylase before starch addition (order 1), mixing TA with pre-gelatinized starch before α-amylase addition (order 2) and co-gelatinizing TA with starch before α-amylase addition (order 3). It was found that the enzyme inhibition was always highest for order 1 because TA could bind with the enzyme active site thoroughly before digestion occurred. Both order 2 and 3 reduced α-amylase inhibition through decreasing binding of TA with the enzyme, which resulted from the non-covalent physical adsorption of TA with gelatinized starch. Interestingly, at low TA concentration, α-amylase inhibition for order 2 was higher than order 3, while at high TA concentration, the inhibition was shown with the opposite trend, which arose from the difference in the adsorption property between the pre-gelatinized and co-gelatinized starch at the corresponding TA concentrations. Moreover, both the crystalline structures and apparent morphology of starch were not significantly altered by TA addition for order 2 and 3. Conclusively, although a polyphenol has an acceptable inhibitory activity in vitro, the actual effect may not reach the expected one when taking processing procedures into account.

scholarly journals Acrolein, an irreversible active-site-directed inhibitor of deoxyribose 5-phosphate aldolase?

1976 ◽  
Vol 153 (2) ◽  
pp. 495-497 ◽  
Author(s):  
D C Wilton
Keyword(s):  
Schiff Base ◽  
Rate Constant ◽  
Active Site ◽  
Order Rate Constant ◽  
Lysine Residue ◽  
Prolonged Incubation ◽  
Enzyme Active Site ◽  
First Order ◽  
Substrate Analogue ◽  
Pseudo First Order

The enzyme deoxyribose 5-phosphate aldolase was irreversibly inactivated by the substrate analogue acrolein with a pseudo-first-order rate constant of 0.324 min-1 and a Ki (apparent) of 2.7 × 10(-4) m. No inactivation was observed after prolonged incubation with the epoxide analogues glycidol phosphate and glycidaldehyde. It is suggested that the acrolein is first activated by forming a Schiff base with the enzyme active-site lysine residue and it is the activated inhibitor that reacts with a suitable-active-site nucleophile.

scholarly journals The Physical Adsorption of Gelatinized Starch With Tannic Acid Decreases the Inhibitory Activity of the Polyphenol Against α-Amylase

2021 ◽  
Author(s):  
Yueyi Wang ◽  
Fangting Bai ◽  
Junwei Cao ◽  
Lijun Sun
Keyword(s):  
Enzyme Inhibition ◽  
Tannic Acid ◽  
Inhibitory Activity ◽  
Adsorption Property ◽  
Physical Adsorption ◽  
Alpha Amylase ◽  
Enzyme Active Site ◽  
Gelatinized Starch ◽  
The Difference

The effects of mixing orders of tannic acid (TA), starch and α-amylase on the enzyme inhibition of TA were studied, including mixing TA with α-amylase before starch addition (order 1), mixing TA with pre-gelatinized starch before α-amylase addition (order 2) and co-gelatinizing TA with starch before α-amylase addition (order 3). It was found that the enzyme inhibition was always highest for order 1 because TA could bind with the enzyme active site thoroughly before digestion occurred. Both order 2 and 3 reduced α-amylase inhibition through decreasing binding of TA with the enzyme, which resulted from the non-covalent physical adsorption of TA with gelatinized starch. Interestingly, at low TA concentration, α-amylase inhibition for order 2 was higher than order 3, while at high TA concentration, the inhibition was shown with opposite trend, which arose from the difference in the adsorption property between the pre-gelatinized and co-gelatinized starch at the corresponding TA concentrations. Besides, both the crystalline structures and apparent morphology of starch were not significantly altered by TA addition for order 2 and 3. Conclusively, although a polyphenol may have an acceptable inhibitory activity in vitro, the actual effect may not reach the expected one when taking processing procedures into account.

scholarly journals Neuraminidase inhibition by chemically sulphated glycopeptides

1979 ◽  
Vol 181 (2) ◽  
pp. 377-385 ◽  
Author(s):  
N Mian ◽  
C E Anderson ◽  
P W Kent
Keyword(s):  
Enzyme Inhibition ◽  
Active Site ◽  
Enzyme Inhibitor ◽  
Helix Pomatia ◽  
Submaxillary Gland ◽  
Competitive Inhibitor ◽  
Duodenal Mucosa ◽  
Dependent Manner ◽  
Enzyme Active Site ◽  
Acetylneuraminic Acid

Chemically sulphated glycopeptides (derived from pig duodenal mucosa) inhibited Clostridium perfringens neuraminidase (EC 3.2.1.18) activity in a pH-dependent manner. Analysis of inhibition kinetics data indicated that, although the enzyme inhibition could not be categorized into any of the classical types of inhibition, it could be interpreted as a function of the size and shape of the substrates used. The enzyme activity was inhibited by 86% and 40% when tested with bovine submaxillary-gland mucin (mol. wt. 4 x 10(5)-40 x 10(5) and N-acetylneuraminyl-lactose (mol. wt. 633) as substrates respectively. Presence of sulphated glycopeptide did not affect the binding of N-acetylneuraminic acid (mol. wt. 309), a competitive inhibitor of Vibrio cholerae neuraminidase, to the enzyme active site. The enzyme inhibition was thus considered to be due to steric hindrance as a consequence of the non-specific interactions between the enzyme molecule and polyanionic sulphated glycopeptide affecting the differential accessibility of the substrate molecules to the enzyme active site. The enzyme-inhibitor interaction could be suppressed by rapid and many-fold dilution of the reaction mixture, by concurrent addition of the inactive enzyme or by partial removal of the sulphate esters from the sulphated glycopeptide molecule by the action of Helix pomatia arylsulphatase (EC 3.1.6.1).

Characterization of an Abnormal Antithrombin (Milano 2) with Defective Thrombin Binding

1986 ◽  
Vol 56 (03) ◽  
pp. 349-352 ◽  
Author(s):  
A Tripodi ◽  
A Krachmalnicoff ◽  
P M Mannucci
Keyword(s):  
Venous Thromboembolism ◽  
Active Site ◽  
Inhibitory Activity ◽  
Antithrombin Iii ◽  
Factor Xa ◽  
Molecular Defect ◽  
Affinity Adsorption ◽  
Italian Family ◽  
Crossed Immunoelectrophoresis

SummaryFour members of an Italian family (two with histories of venous thromboembolism) had a qualitative defect of antithrombin III reflected by normal antigen concentrations and halfnormal antithrombin activity with or without heparin. Anti-factor Xa activities were consistently borderline low (about 70% of normal). For the propositus’ plasma and serum the patterns of antithrombin III in crossed-immunoelectrophoresis with or without heparin were indistinguishable from those of normal plasma or serum. A normal affinity of antithrombin III for heparin was documented by heparin-sepharose chromatography. Affinity adsorption of the propositus’ plasma to human α-thrombin immobilized on sepharose beads revealed defective binding of the anti thrombin III to thrombin-sepharose. Hence the molecular defect of this variant appears to be at the active site responsible for binding and neutralizing thrombin, thus accounting for the low thrombin inhibitory activity.

scholarly journals Conformation-Specific Inhibitory Anti-MMP-7 Monoclonal Antibody Sensitizes Pancreatic Ductal Adenocarcinoma Cells to Chemotherapeutic Cell Kill

Cancers ◽  
2021 ◽  
Vol 13 (7) ◽  
pp. 1679
Author(s):  
Vishnu Mohan ◽  
Jean P. Gaffney ◽  
Inna Solomonov ◽  
Maxim Levin ◽  
Mordehay Klepfish ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Active Site ◽  
Drug Targets ◽  
Fas Ligand ◽  
Specific Activity ◽  
Matrix Metalloproteases ◽  
Ductal Adenocarcinoma ◽  
Enzyme Active Site ◽  
Induced Apoptosis

Matrix metalloproteases (MMPs) undergo post-translational modifications including pro-domain shedding. The activated forms of these enzymes are effective drug targets, but generating potent biological inhibitors against them remains challenging. We report the generation of anti-MMP-7 inhibitory monoclonal antibody (GSM-192), using an alternating immunization strategy with an active site mimicry antigen and the activated enzyme. Our protocol yielded highly selective anti-MMP-7 monoclonal antibody, which specifically inhibits MMP-7′s enzyme activity with high affinity (IC50 = 132 ± 10 nM). The atomic model of the MMP-7-GSM-192 Fab complex exhibited antibody binding to unique epitopes at the rim of the enzyme active site, sterically preventing entry of substrates into the catalytic cleft. In human PDAC biopsies, tissue staining with GSM-192 showed characteristic spatial distribution of activated MMP-7. Treatment with GSM-192 in vitro induced apoptosis via stabilization of cell surface Fas ligand and retarded cell migration. Co-treatment with GSM-192 and chemotherapeutics, gemcitabine and oxaliplatin elicited a synergistic effect. Our data illustrate the advantage of precisely targeting catalytic MMP-7 mediated disease specific activity.

scholarly journals 1250. Novel Boronic Acid Transition State Analogs (BATSI) with in vitro inhibitory activity against class A, B and C β-lactamases

2020 ◽  
Vol 7 (Supplement_1) ◽  
pp. S643-S643
Author(s):  
Maria F Mojica ◽  
Christopher Bethel ◽  
Emilia Caselli ◽  
Magdalena A Taracila ◽  
Fabio Prati ◽  
...  
Keyword(s):  
Active Site ◽  
Inhibitory Activity ◽  
Covalent Bond ◽  
Thiol Group ◽  
Viable Cell ◽  
Free Thiol ◽  
Transition State Analogs ◽  
Cell Counts ◽  
Free Thiol Group

Abstract Background Catalytic mechanisms of serine β-lactamases (SBL; classes A, C and D) and metallo-β-lactamases (MBLs) have directed divergent strategies towards inhibitor design. SBL inhibitors act as high affinity substrates that -as in BATSIs- form a reversible, dative covalent bond with the conserved active site Ser. MBL inhibitors bind the active-site Zn2+ ions and displace the nucleophilic OH-. Herein, we explore the efficacy of a series of BATSI compounds with a free-thiol group at inhibiting both SBL and MBL. Methods Exploratory compounds were synthesized using stereoselective homologation of (+) pinandiol boronates to introduce the amino group on the boron-bearing carbon atom, which was subsequently acylated with mercaptopropanoic acid. Representative SBL (KPC-2, ADC-7, PDC-3 and OXA-23) and MBL (IMP-1, NDM-1 and VIM-2) were purified and used for the kinetic characterization of the BATSIs. In vitro activity was evaluated by a modified time-kill curve assay, using SBL and MBL-producing strains. Results Kinetic assays revealed that IC50 values ranged from 1.3 µM to >100 µM for this series. The best compound, s08033, demonstrated inhibitory activity against KPC-2, VIM-2, ADC-7 and PDC-3, with IC50 in the low μM range. Reduction of at least 1.5 log10-fold of viable cell counts upon exposure to sub-lethal concentrations of antibiotics (AB) + s08033, compared to the cells exposed to AB alone, demonstrated the microbiological activity of this novel compound against SBL- and MBL-producing E. coli (Table 1). Table 1 Conclusion Addition of a free-thiol group to the BATSI scaffold increases the range of these compounds resulting in a broad-spectrum inhibitor toward clinically important carbapenemases and cephalosporinases. Disclosures Robert A. Bonomo, MD, Entasis, Merck, Venatorx (Research Grant or Support)

scholarly journals Toward the Identification of Potential α-Ketoamide Covalent Inhibitors for SARS-CoV-2 Main Protease: Fragment-Based Drug Design and MM-PBSA Calculations

Processes ◽  
2021 ◽  
Vol 9 (6) ◽  
pp. 1004
Author(s):  
Mahmoud A. El Hassab ◽  
Mohamed Fares ◽  
Mohammed K. Abdel-Hamid Amin ◽  
Sara T. Al-Rashood ◽  
Amal Alharbi ◽  
...  
Keyword(s):  
Drug Design ◽  
Active Site ◽  
Binding Free Energy ◽  
Stable Complex ◽  
Active Site Residues ◽  
Structure Based Drug Design ◽  
Enzyme Active Site ◽  
Potential Inhibitor ◽  
Main Protease ◽  
Covalent Docking

Since December 2019, the world has been facing the outbreak of the SARS-CoV-2 pandemic that has infected more than 149 million and killed 3.1 million people by 27 April 2021, according to WHO statistics. Safety measures and precautions taken by many countries seem insufficient, especially with no specific approved drugs against the virus. This has created an urgent need to fast track the development of new medication against the virus in order to alleviate the problem and meet public expectations. The SARS-CoV-2 3CL main protease (Mpro) is one of the most attractive targets in the virus life cycle, which is responsible for the processing of the viral polyprotein and is a key for the ribosomal translation of the SARS-CoV-2 genome. In this work, we targeted this enzyme through a structure-based drug design (SBDD) protocol, which aimed at the design of a new potential inhibitor for Mpro. The protocol involves three major steps: fragment-based drug design (FBDD), covalent docking and molecular dynamics (MD) simulation with the calculation of the designed molecule binding free energy at a high level of theory. The FBDD step identified five molecular fragments, which were linked via a suitable carbon linker, to construct our designed compound RMH148. The mode of binding and initial interactions between RMH148 and the enzyme active site was established in the second step of our protocol via covalent docking. The final step involved the use of MD simulations to test for the stability of the docked RMH148 into the Mpro active site and included precise calculations for potential interactions with active site residues and binding free energies. The results introduced RMH148 as a potential inhibitor for the SARS-CoV-2 Mpro enzyme, which was able to achieve various interactions with the enzyme and forms a highly stable complex at the active site even better than the co-crystalized reference.

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