Current Enzyme Inhibition - Volume 17, Issue 3, 2021

Despite the significant number of irreversible inhibitors developed over the years, strong prejudices remain for this type of therapeutic molecule, particularly in the area of drug development. New generations of covalent targeted inhibitors are, however, in development, and interest is increasingly growing. In fact, the new generation of covalent inhibitors has a weakly reactive species (warhead) that is able, in a particular context, to selectively form a chemical bond with a given amino acid residue, which can be irreversible or reversible. In addition to new selective warheads, new amino acids are also targeted. In the following text, we will focus on covalent targeted inhibitors that selectively alkylate histidine.

Background: Overactivity of fibroblast growth factor receptor 1 (FGFR1) is associated with various tumors, particularly breast cancer, prostate cancer, non-small-cell lung carcinoma, myeloproliferative diseases, which makes this protein kinase a promising therapeutic target for anticancer therapy. Objective: The main aim of this study is to identify novel FGFR1 inhibitors. Methods: In order to find FGFR1 inhibitors, virtual screening experiments were performed using AutoDock software. Best-scored compounds were tested in vitro using P32 radioactive kinase assay. Results: Small-molecular inhibitors of protein kinase FGFR1 were identified among indazole derivatives. The most active compound [3-(3,4-dichloro-phenyl)-1H-indazol-5-yl]-(2,3-dihydro-1Hcyclopenta[ b]quinolin-9-yl)-amine (1) inhibits FGFR1 with IC50 value of 100 nM. According to molecular docking results, this compound interacts simultaneously with adenine- and phosphate-binding regions of protein kinase FGFR1. The structure-activity relationships have been investigated and binding mode has been predicted. Conclusion: Compound 1 can be used for further structural optimization and biological research.

Background and Aims: This study aims to alleviate the microbiologically affected corrosion that occurred by sulfate-reducing bacteria (SRB) through synthesizing a bio-based polyurethane polymer and its nanocomposite coating, silver nanoparticles (PU-Ag). Moreover, it will also evaluate the effect of PU alone and PU-Ag as inhibitors for adenylyl sulfate reductase (APS), the main enzyme for sulfate reduction. Methods: In this study, the PU was prepared from vegetable soybean oil, and the silver nanoparticles (Ag-NPs) with a concentration of 1% were coated to the PU forming a nanocomposite. The PU and the PU-Ag were characterized and evaluated as inhibitors of the APS reductase enzyme. Results: The results obtained from FTIR, UV, DLS, TEM, and XRD confirmed the preparation structure of the PU and PU-Ag. Furthermore, the PU/PU-Ag competitively inhibited the APS reductase with an inhibition constant equal to 35.7 and 11 mg, respectively. These indicated the exerted inhibitory effect of PU/PU-Ag upon the activity of the APS reductase enzyme. Conclusion: The APS reductase enzyme produced by SRB, which is recorded as a big problem in the oil and gas industry such as pitting corrosion of tanks and pipelines, could be inhibited by PU and PU-Ag.

Background: Plants are the main source of drugs for the therapy of a large number of diseases. Objective: The aim of the present work is to evaluate the in vitro antioxidant and anti-α-amylase and anti- peroxidases (HPR) potentials of phenolic extracts obtained from three spontaneous plants growing in the South of Algeria, namely Chamomilla pubescens, Pulicaria crispa, and Rhanterium adpressum. This is the first report on the study of α-amylase and horseradish peroxidase (HRP) inhibitory activity for phenolic extracts from the Chamomilla pubescens and Pulicaria crispa plants. Methods: The antioxidant activity was evaluated in vitro using four tests: DPPH, CUPRAC, FRAP, and ABTS. The phenolic, flavonoid, and tannin compounds of the three selected Algerian plants were quantified. Also, the inhibition of α-amylase and HRP was evaluated. Results: The quantification of the total phenolic contents revealed that they are variable and widely dependent on extraction solvents. The highest content was recorded by the ethyl acetate extract of Chamomilla pubescens (flowers) 774.93±60.14mg/100 g of dry matter. In all the antioxidant tests, ethyl acetate extracts showed the most effective activity, of which the best was (VCEAC = 65.62 ±0.50 μM/g dry matter) of Pulicaria crispa for the DPPH test. Furthermore, the results of α-amylase and peroxidase inhibitory effects indicated that all plants extracts have inhibitory effects on the two enzymes, with AEIC values ranging from 76.55±3.54 to 149.54±6.68 μM/g of dry matter for the α-amylase and CEIC values ranging from 8.89±2.22 to 9668.31±254.42 μM/g of dry matter for the peroxidase (HRP). Conclusion: The present study results suggest that the three Algerian spontaneous plant species (Rhanterium adpressum, Pulicaria crispa and Chamomilla pubescens) inhibit peroxidase and α-amylase and exhibit a high antioxidant activity which can be related to the treatment of diabetes and thyroid diseases.

Background: The amino acid sequence of an α-amylase of the yeast Saccharomycopsis fibuligera R64 (SfamyR64) contains two putative N-linked glycosylation sites N153 and N224. N224 is hypothetically responsible for the binding of starch substrate because it is highly conserved among SfamyR64 homologs. Objective: To test whether N224 plays a key role in enzyme activity and stability. Methods: N224Q substitution was introduced by site-directed mutagenesis. The wild type and the mutant were independently over-produced in Pichia pastoris KM71. Activities of the wild type and of the mutant were compared, and their thermal-stability was assessed using heat treatments. The evolutionary relationship of SfamyR64 with its structural homologs with different glycosylation patterns was examined. Results: Activity of the N224Q mutant was approximately 80% lower than that of the wild type. The mutant showed no activity after 10 min of pre-incubation at 50 °C, whereas the wild type SfamyR64 showed activity until 30 min of treatment. Sfamy appeared to have evolved earlier than its structural homolog. Conclusion: SfamyR64 N224 is crucial for enzyme activity and thermal stability. This glycosylation site is unique for fungal and bacterial α-amylases.

Background: Garlic, being a well-known medicinal plant, is the most commonly used culinary spice worldwide. Investigation of protease inhibitor isolated from garlic leads to a promising contender in pharmacognostic and pharmacological studies. Objective/Introduction: Protease Inhibitor (PI) from 'garlic' (Allium sativum) was analyzed for its biological role as an antioxidant, antimicrobial, and anti-inflammatory agent. Methods: Antioxidant activity was evaluated using ferric ion reducing antioxidant power (FRAP) and 2, 2-diphenyl-1-picrylhydrazyl (DPPH) assays. The anti-inflammatory activity was assessed using trypsin inhibitory assay and heat-induced albumin denaturation method. The antimicrobial activity was examined in broth against E. coli and B. Subtilis. The crystallization was setup using the hanging drop method. Results: ASPI showed DPPH radical scavenging with IC50 values 561±0.337 μg/ml. Also, ASPI showed the highest value of 0.699±0.009 mM at 1000 μg/ml and the lowest i.e. 0.181±0.006 mM at 100 μg/ml in FRAP assay. Ascorbic acid was taken as standard in both cases. ASPI showed IC50 values of 651±0.532 μg/ml and ~657±1.802 μg/ml, respectively. The antibacterial role of ASPI was testified, and results showed maximum inhibition against E. coli (ATCC 25922) i.e., 87.8 ±0.602% but no inhibition against B. subtilis (MTCC 736). Cuboidal shaped crystals of the ASPI were obtained in 4-6 weeks using 0.2M calcium chloride dihydrate, 0.1M sodium acetate trihydrate, 20 % isopropanol. Conclusion: ASPI has tremendous potential in the development of suitable drugs in pharmaceutical industries against diseases due to the generation of reactive oxygen species and cancer. The cuboidal crystals were obtained, which is the first study in the context of crystallization of ASPI to date.

Background: A series of bis(indolyl)methanes (3a-3o) have been synthesized using a greener and new approach using the reaction of different substituted aldehydes and indole in the presence of an easily available and biodegradable base such as piperidine in acetic acid at room temperature and characterized with UV (Ultraviolet-visible spectroscopy), Gas Chromatography-Mass Spectrometry (GCMS), Proton Nuclear Magnetic Resonance (H-NMR), and Fourier Transform Infrared Spectroscopy (FTIR). Methods: All 15 newly synthesized compounds (3a-3o) were subjected to in-vitro anti-microbial activity determination and compared with the known standard drug ciprofloxacin (1-2 μg/mL). Our in-silico analysis on the target protein, pdb id: 1d7u suggested that these analogues would be highly active against bacterial targets and thus, would act as good antimicrobial agents. Results: All 15 newly synthesized compounds (3a-3o) displayed potent activity on various experimental microbial strains (1.0-1.4 μg/mL). Compound, 3k was obtained as the best docked compound against common bacterial target enzyme, (pdb id:1d7u). The standard, Ciprofloxacin, retained the docking score of -111.3 Kcal/mol with similar binding amino acid residues (LYS272 (Pi-cation); ALA A:245 (Pisigma); TRP A:138 (Pi-Pi); ALA A:112; and MET A:141 (Pi-alkyl)) as of inbound. Conclusion: We believe that our current study would shed more light on the development of potent bis(indolyl)methanes as antimicrobial agents.

Background: Obesity is a growing global health problem. Obesity leads to cardiovascular disorders, musculoskeletal disorders, diabetes, and certain types of cancer. One of the approaches to controlling and treating obesity has involved inhibition of dietary lipid digestion by pancreatic lipase inhibitors. Microbes and plant sources provide a rich source of enzyme inhibitors, including pancreatic lipase inhibitors that can be developed as a drug for obesity treatment. Objective: The objective of the work, mainly focuses and highlights the isolation of actinomycetes and screening for pancreatic lipase inhibitors production. Methods: Actinomycetes were isolated from soil samples by pre-treatment of samples and using selective growth medium with and without antibiotics. Isolated actinomycetes were grown in fermentation conditions and metabolites were extracted with isopropyl alcohol and the solvent evaporated to get crude material. Extract of each isolate screened for pancreatic lipase inhibition using p- nitrophenyl palmitate as substrate by spectroscopic method. Results: A total of 86 actinomycetes strains were isolated from soil samples. Out of 86 extracts,10 extracts have shown positive results for porcine pancreatic lipase inhibition, and inhibition activity was in the range of 10-80%. 50 % inhibitory concentration determined using 1 to 8 mg/mL extract in the assay. Extract of isolate A9, B3, and C6 having 50 % inhibitory activity below 3 mg/mL concentration and Orlistat as a standard has shown 50 % inhibitory activity at below 1 mg/mL concentration. Conclusion: The results conclude that actinomycetes are potential source of lipase inhibitors, which may lead to valuable novel drugs for obesity treatment.

Background: Inhibition of α-amylase and α-glucosidase is considered as an important therapeutic target to manage type 2 diabetes mellitus (T2DM), reducing postprandial hyperglycemia (PPHG). Objective: The present work explored the antidiabetic activities of five artificial food colorings by α- amylase and α-glucosidase enzyme inhibition in vitro and in Silico. Methods: In this study, inhibition of α-amylase and α-glucosidase was evaluated. Further, the interaction between enzymes (α-amylase and α-glucosidase) and ligands (food colorings) was followed by QSAR and molecular docking studies. Results: The in vitro results obtained show that the blue patent (SIN131) exhibited more potent inhibition with IC50 values of 0.03± 0.01 mM and 0.014±0.001 mM against α-amylase and α-glucosidase inhibition respectively compared to acarbose. The QSAR study found a strong correlation between IC50 values with four molecular descriptors. This linear regression confirms that a strong polarity (Apol) and low hydrophobia (ALogP) favor the inhibitory effect of these colorings toward both enzymes. Also, a negative role of the number of heavy atoms has been demonstrated in the phenomenon of inhibition of this enzyme. Finally, the descriptor lumo (electronic affinity) plays a crucial role on the inhibitory power of these dyes toward both enzymes by electron transfer. The virtual screening of the inhibition of α-amylase and α-glucosidase by these colorings, using Molegro Virtual Docker (MVD), allowed us to obtain stable complexes with interaction energies resulting from the place of hydrogen bonds and several hydrophobic interactions. However, the sulfonate groups of these colorings can be the major factors in the inhibition of these enzymes. On the other hand, Rerank Score with the pose are perfectly correlated (R2> 0.76) to the inhibitory activity of these food colorings measured experimentally. Conclusion: The present study suggests that the Blue Patent V (SIN131) effectively acts as α-amylase and α-glucosidase inhibitor leading to a reduction in starch hydrolysis and eventually to lower glucose levels.