Deciphering the role of aquaporin 1 in the adaptation of the stinging catfish Heteropneustes fossilis to environmental hypertonicity by molecular dynamics simulation studies.

A thorough investigation of the water permeability of H. fossilis aquaporin 1 (hfAQP1) in a hypertonic environment can provide a useful insight into the understanding of the underlying molecular mechanism of its high tolerance to salinity. Here, we constructed a 3 D homology model of hfAQP1 by taking Bos taurus AQP1, AQP0, and human AQP2 as templates using I-TASSER. The model obtained has similar structural organizations with mammalian AQP1s in all aspects. We investigated the water permeability of the modeled hfAQP1 in a 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) membrane under neutral and 100 mM hypersalinity by subjecting each system to a 100 ns molecular dynamics simulation. Our results show that hypersalinity hinders water permeation across the membrane through the hfAQP1 channel. A change in the intermolecular distance between key residues of the ar/R selectivity filter along with charge redistribution resulted in the accommodation of only 2-6 water molecules inside the channel at once under hypersaline conditions. We investigated the mRNA expression pattern of hfaqp1 in osmoregulatory organs of H. fossilis in response to 100 mM hypertonicity by using qPCR analysis. The transcript was downregulated in kidney and GI tract, but upregulated in the Gills. Thus, the catfish survive in a hypertonic environment by reducing the transport of water in its cellular systems and downregulating the expression of the hfaqp1 gene. The results observed in our study can shed more light on the functionality of AQP1 in catfishes under salinity stress and aid in future researches on solving more gating mechanisms involved in its regulation.Communicated by Ramaswamy H. Sarma.

Understanding the binding mechanism for potential inhibition of SARS-CoV-2 Mpro and exploring the modes of ACE2 inhibition by hydroxychloroquine.

As per the World Health Organization report, around 226 844 344 confirmed positive cases and 4 666 334 deaths are reported till September 17, 2021 due to the recent viral outbreak. A novel coronavirus (severe acute respiratory syndrome coronavirus 2 [SARS-CoV-2]) is responsible for the associated coronavirus disease (COVID-19), which causes serious or even fatal respiratory tract infection and yet no approved therapeutics or effective treatment is currently available to combat the outbreak. Due to the emergency, the drug repurposing approach is being explored for COVID-19. In this study, we attempt to understand the potential mechanism and also the effect of the approved antiviral drugs against the SARS-CoV-2 main protease (Mpro). To understand the mechanism of inhibition of the malaria drug hydroxychloroquine (HCQ) against SARS-CoV-2, we performed molecular interaction studies. The studies revealed that HCQ docked at the active site of the Human ACE2 receptor as a possible way of inhibition. Our in silico analysis revealed that the three drugs Lopinavir, Ritonavir, and Remdesivir showed interaction with the active site residues of Mpro. During molecular dynamics simulation, based on the binding free energy contributions, Lopinavir showed better results than Ritonavir and Remdesivir.

Peptidomics and proteogenomics: background, challenges and future needs.

INTRODUCTION: With available genomic data and related information, it is becoming possible to better highlight mutations or genomic alterations associated with a particular disease or disorder. The advent of high-throughput sequencing technologies has greatly advanced diagnostics, prognostics, and drug development. AREAS COVERED: Peptidomics and proteogenomics are the two post-genomic technologies that enable the simultaneous study of peptides and proteins/transcripts/genes. Both technologies add a remarkably large amount of data to the pool of information on various peptides associated with gene mutations or genome remodeling. Literature search was performed in the PubMed database and is up to date. EXPERT OPINION: This article lists various techniques used for peptidomic and proteogenomic analyses. It also explains various bioinformatics workflows developed to understand differentially expressed peptides/proteins and their role in disease pathogenesis. Their role in deciphering disease pathways, cancer research, and biomarker discovery using biofluids is highlighted. Finally, the challenges and future requirements to overcome the current limitations for their effective clinical use are also discussed.

Proteomics and Machine Learning Approaches Reveal a Set of Prognostic Markers for COVID-19 Severity With Drug Repurposing Potential.

The pestilential pathogen SARS-CoV-2 has led to a seemingly ceaseless pandemic of COVID-19. The healthcare sector is under a tremendous burden, thus necessitating the prognosis of COVID-19 severity. This in-depth study of plasma proteome alteration provides insights into the host physiological response towards the infection and also reveals the potential prognostic markers of the disease. Using label-free quantitative proteomics, we performed deep plasma proteome analysis in a cohort of 71 patients (20 COVID-19 negative, 18 COVID-19 non-severe, and 33 severe) to understand the disease dynamics. Of the 1200 proteins detected in the patient plasma, 38 proteins were identified to be differentially expressed between non-severe and severe groups. The altered plasma proteome revealed significant dysregulation in the pathways related to peptidase activity, regulated exocytosis, blood coagulation, complement activation, leukocyte activation involved in immune response, and response to glucocorticoid biological processes in severe cases of SARS-CoV-2 infection. Furthermore, we employed supervised machine learning (ML) approaches using a linear support vector machine model to identify the classifiers of patients with non-severe and severe COVID-19. The model used a selected panel of 20 proteins and classified the samples based on the severity with a classification accuracy of 0.84. Putative biomarkers such as angiotensinogen and SERPING1 and ML-derived classifiers including the apolipoprotein B, SERPINA3, and fibrinogen gamma chain were validated by targeted mass spectrometry-based multiple reaction monitoring (MRM) assays. We also employed an in silico screening approach against the identified target proteins for the therapeutic management of COVID-19. We shortlisted two FDA-approved drugs, namely, selinexor and ponatinib, which showed the potential of being repurposed for COVID-19 therapeutics. Overall, this is the first most comprehensive plasma proteome investigation of COVID-19 patients from the Indian population, and provides a set of potential biomarkers for the disease severity progression and targets for therapeutic interventions.

Proteomic investigation reveals dominant alterations of neutrophil degranulation and mRNA translation pathways in patients with COVID-19.

The altered molecular proteins and pathways in response to COVID-19 infection are still unclear. Here, we performed a comprehensive proteomics-based investigation of nasopharyngeal swab samples from patients with COVID-19 to study the host response by employing simple extraction strategies. Few of the host proteins such as interleukin-6, L-lactate dehydrogenase, C-reactive protein, Ferritin, and aspartate aminotransferase were found to be upregulated only in COVID-19-positive patients using targeted multiple reaction monitoring studies. The most important pathways identified by enrichment analysis were neutrophil degranulation, interleukin-12 signaling pathways, and mRNA translation of proteins thus providing the detailed investigation of host response in COVID-19 infection. Thus, we conclude that mass spectrometry-detected host proteins have a potential for disease severity progression; however, suitable validation strategies should be deployed for the clinical translation. Furthermore, the in silico docking of potential drugs with host proteins involved in the interleukin-12 signaling pathway might aid in COVID-19 therapeutic interventions.

Understanding the binding mechanism of succinic acid against phospholipase A2 from bee venom.

Phospholipase A2 (PLA2 ) is responsible for the release of fatty acids from glycerophospholipids. PLA2 is commonly found in mammalian tissues. It is also found in venom from different animals ranging from insects, arachnid, and snakes. The release of arachidonic acid in large amount results in inflammation and pain. Identification of compounds that can inhibit the activity of PLA2 is of large scientific and medicinal interest as these compounds can act as antidotes toward snake bites and bee stings. Among the different compounds that have been tested for inhibition of PLA2 , a secondary metabolite succinic acid is identified to inhibit PLA2 activity. The inhibition was analyzed using an in vitro PLA2 inhibition assay and isothermal titration calorimetry (ITC) studies. The molecular mechanism of the mode of inhibition was studied using molecular docking and simulation studies.

Serine protease from Tricosanthus tricuspidata accelerates healing of Echis carinatus venom-induced necrotic wound.

Echis carinatus (EC) envenomation causes severe immune response by the accumulation of tissue debris in the form of DAMPs resulting in chronic inflammation and progressive tissue necrosis at the bitten site. Clearing of tissue debris is a prerequisite to enhance the healing of venom-induced necrotic wounds. Tricosanthus tricuspidata is a medicinal plant used extensively for the treatment of snake bite-induced toxicities. The active component responsible for the observed pharmacological action is a serine protease, tricuspidin. The topical application of tricuspidin was able to neutralize ECV-induced mouse footpad tissue necrosis and open wound in rabbits. Tricuspidin exerted its healing action via proteolytic activity as a consequence of upregulation of MMP-8 and down regulation of MMP-9. Further, tricuspidin reduced ECV-induced inflammation by decreasing the expression of TNF-α, IL-6 and MPO, and by increasing the level of VEGF-A and TGF-ß1. The modulation of ECV induced immune/inflammatory mediators by tricuspidin was found to be more effective than trypsin. Moreover, tricuspidin and trypsin activated MAPKs via protease activated receptors-2 (PAR-2). These data indicate that the proteolytic activity of tricuspidin directly involved in the healing of ECV-induced chronic wound.

Combinatorial inhibition of Angiotensin converting enzyme, Neutral endopeptidase and Aminopeptidase N by N-methylated peptides alleviates blood pressure and fibrosis in rat model of dexamethasone-induced hypertension.

Angiotensin converting enzyme (ACE), neutral endopeptidase (NEP) and aminopeptidase N (APN) are responsible for generation of vasoactive peptides that regulates vasoconstriction, vasodilation and natriuresis, which altogether regulate blood pressure. Cumulative inhibition of ACE, NEP and APN effectively blocks the progression of respective pathways. In this study, N-methylated peptide inhibitors F-N(Me)H-L, V-N(Me)F-R and R-N(Me)V-Y were synthesized against ACE, NEP and APN respectively, using their respective physiological substrates. F-N(Me)H-L inhibited ACE activity with an IC50 of 83 nmol/L, V-N(Me)F-R inhibited NEP activity with an IC50 of 1.173 µmol/L and R-N(Me)V-Y inhibited APN activity with an IC50 of 3.94 nmol/L respectively. Further, the anti-hypertensive effect of N-methylated peptides was evaluated using rat model of dexamethasone-induced hypertension. Individual peptides and their cocktail treatment were started from day 6 of the study period and blood pressure was measured on every alternate day during 15 day study. Administration of F-N(Me)H-L (138 ± 3 mmHg) and cocktail of all the three peptides at a dose of 100 mg/kg significantly reduced systolic blood pressure (SBP) compared to dexamethasone group (SBP of Groups-dexamethasone; (167 ± 5 mmHg), F-N(Me)H-L (138 ± 3 mmHg), and Cocktail (122 ± 3 mmHg). Anti-hypertensive, anti-hypertrophic and anti-fibrotic effects of N-methylated peptides and cocktail was further reflected by the decreased levels of circulating Ang II and increased ANP levels in sera of hypertensive rats along with decrease in collagen deposition in heart and kidney. Though, ACE inhibition is adequate to reduce SBP, targeting NEP and APN along with ACE is beneficial in tackling hypertension and associated fibrosis of heart.

Nutritional Status of Adolescent Girls Belonging to the Tea Garden Estates of Sivasagar District, Assam, India.

BACKGROUND: Any deficiency or inadequate dietary pattern can lead to poor nutrition which can further influence both growth and development throughout from infancy to adolescence. Since adolescents represent the next generation of parents, it is important to monitor their nutritional status at this crucial stage. Thus, this study aimed to explore the factors associated with nutritional status among adolescent girls belonging to these tea gardens. OBJECTIVE: The objective of this community-based cross-sectional study was to assess the nutritional status of adolescent girls belonging to the tea garden community and the association of the sociodemographic factors with it. MATERIALS AND METHODS: Anthropometric measurement was taken among adolescent girls in the tea estates of Nazira subdivision of Sivasagar district, Assam. The pattern of dietary intake among adolescents was also studied. The statistical analysis was done using SPSS version 15. RESULTS: The prevalence of thinness and stunting across 265 adolescent girls was 49.4% and 50.6%, respectively. Calorie and protein deficits were found to be 76.60% and 65%, respectively. Majority of the respondents, i.e., 66.80% of the participants, had a poor intake of essential food constituents. Moreover, 76.21% of the respondents were anemic. The association of different sociodemographic factors with thinness, inadequate protein intake, and anemia were found during the study. CONCLUSION: Thinness and stunting along with protein-energy malnutrition and inadequate intake of important food groups were prevalent in adolescent tea community girls. Overall, the public health burden of malnutrition is still a persisting health problem in the tea gardens of Assam.

Inhibitory effects of ascorbic acid toward snake venom metalloproteinase (SVMP) from Indian Echis carinatus venom: Insights from molecular modeling and binding studies.

Classical antivenom therapy is unable to shield complications of viper bite and has limitations such as anaphylaxis and serum sickness. Snake venom metalloproteinases are responsible for local tissue damage and hemorrhage at the bitten site in viper envenomation, and this has led to a persistent search for metalloproteinase inhibitors. Here, we report the inhibitory effects of ascorbic acid against metalloproteinase from Echis carinatus venom both in-silico and in-vitro. Ascorbic acid effectively inhibited the proteolytic activity of E. carinatus venom in a dose-dependent manner. Interaction studies of ascorbic acid with purified ecarin using isothermal titration calorimetry showed favorable binding energy and energetics. The molecular docking of ascorbic acid with ecarin revealed important interactions with residues at the active site pocket of ecarin. It was observed that the ligand behaves as a chelating inhibitor. Thus, the backbone structural scaffold of ascorbic acid can find potential use as building blocks in designing drug-like molecules for viper bite management.

Crystal structures and Hirshfeld surface analyses of 2-[(4,6-di-amino-pyrimidin-2-yl)sulfan-yl]-N-(pyridin-2-yl)acetamide and 2-[(4,6-di-amino-pyrimidin-2-yl)sulfan-yl]-N-(pyrazin-2-yl)acetamide.

In the title compounds, C11H12N6OS (I) and C10H11N7OS (II), the di-amino-pyrimidine ring makes dihedral angles of 71.10 (9)° with the pyridine ring in (I) and 62.93 (15)° with the pyrazine ring in (II). The ethanamine group, -CH2-C(=O)-NH- lies in the plane of the pyridine and pyrazine rings in compounds (I) and (II), respectively. In both compounds, there is an intra-molecular N-H⋯N hydrogen bond forming an S(7) ring motif and a short C-H⋯O inter-action forming an S(6) loop. In the crystals of both compounds, mol-ecules are linked by pairs of N-H⋯N hydrogen bonds, forming inversion dimers with R22(8) ring motifs. In (I), the dimers are linked by N-H⋯O and N-H⋯N hydrogen bonds, forming layers parallel to (1[Formula: see text] [Formula: see text]). The layers are linked by offset π-π inter-actions [inter-centroid distance = 3.777 (1) Å], forming a three-dimensional supra-molecular structure. In (II), the dimers are linked by N-H⋯O, N-H⋯N and C-H⋯O hydrogen bonds, also forming a three-dimensional supra-molecular structure.

Orphan Three-Finger Toxins Bind at Tissue Factor-Factor VIIa Interface to Inhibit Factor X Activation: Identification of Functional Site by Docking.

Three-finger toxins (3FTxs) contribute to toxicity of venomous snakes belonging to the family Elapidae. Currently, functions of a considerable proportion of 3FTxs are still unknown. Here, we describe the function of orphan group I 3FTxs consisting of four members. We also identified a new member of this group by sequencing a transcript isolated from Naja naja venom. This transcript, named najalexin, is identical to that previously described 3FTx from Naja atra venom gland, and shared high sequence identity with ringhalexin from Hemachatus haemachatus and a hypothetical protein from Ophiophagus hannah (here named as ophiolexin). The three-dimensional structure, as predicted by molecular modeling, showed that najalexin and ophiolexin share the same conserved structural organization as ringhalexin and other 3FTxs. Since ringhalexin inhibits the activation of factor X by the tissue factor-factor VIIa complex (TF-FVIIa), we evaluated the interaction of this group of 3FTxs with all components using in silico protein-protein docking studies. The binding of orphan group I 3FTxs to TF-FVIIa complex appears to be driven by their interaction with TF. They bind to fibronectin domain closer to the 170-loop of the FVIIa heavy chain to inhibit factor X activation. The docking studies reveal that functional site residues Tyr7, Lys9, Glu12, Lys26, Arg34, Leu35, Arg40, Val55, Asp56, Cys57, Cys58, and Arg65 on these 3FTxs are crucial for interaction. In silico replacement of these residues by Ala resulted in significant effects in the binding energies. Furthermore, these functional residues are not found in other groups of 3FTxs, which exhibit distinct pharmacological properties.

EC-PIII, a novel non-hemorrhagic procoagulant metalloproteinase: Purification and characterization from Indian Echis carinatus venom.

Procoagulant snake venom toxins find extensive use as reagents in laboratory tests and diagnostic kits. In the present study we report a novel P-III class procoagulant SVMP, EC-PIII from Echis carinatus venom. EC-PIII was purified using a combination of gel-filtration and anion-exchange chromatography. It has a molecular mass of 110kDa and is a dimeric protein as determined by SDS-PAGE. DLS results show that the protein is homogenous and stable in solution. Peptide mass fingerprinting revealed that the peptides obtained show high homology to the other members of SVMP family. The enzymatic studies revealed that EC-PIII shows protease activity and is inhibited by metalloproteinase inhibitors such as EDTA. EC-PIII exhibits procoagulant effect under in-vitro conditions. Local toxicity studies revealed that EC-PIII is devoid of hemorrhagic as well as myotoxic activities. This is the first report of a non-hemorrhagic SVMP to be identified from Indian Echis carinatus venom. EC-PIII can find potential use in diagnostic and other therapeutic uses owing to its biochemical and pharmacological properties.

Crystal structures of 2-[(4,6-di-amino-pyrimidin-2-yl)sulfan-yl]-N-(2,4-di-methyl-phen-yl)acetamide and 2-[(4,6-di-amino-pyrimidin-2-yl)sulfan-yl]-N-(3-meth-oxy-phen-yl)acetamide.

In the title compounds, C14H17N5OS (I) and C13H15N5O2S (II), the dihedral angle between the pyrimidine and benzene rings is 58.64 (8)° in (I) and 78.33 (9)° in (II). In both compounds, there is an intra-molecular C-H⋯O hydrogen bond, and in (II) there is also an intra-molecular N-H⋯N hydrogen bond present. In the crystals of both compounds, a pair of N-H⋯N hydrogen bonds links the individual mol-ecules to form inversion dimers with R22(8) ring motifs. In (I), the dimers are linked by N-H⋯O and C-H⋯O hydrogen bonds, enclosing R21(14), R21(11) and R21(7) ring motifs, forming layers parallel to the (100) plane. There is also an N-H⋯π inter-action present within the layer. In (II), the inversion dimers are linked by N-H⋯O hydrogen bonds enclosing an R44(18) ring motif. The presence of N-H⋯O and C-H⋯O hydrogen bonds generate an R21(6) ring motif. The combination of these various hydrogen bonds results in the formation of layers parallel to the (1-11) plane.

Comparison of proteomic profiles of the venoms of two of the 'Big Four' snakes of India, the Indian cobra (Naja naja) and the common krait (Bungarus caeruleus), and analyses of their toxins.

Snake venoms are mixtures of biologically-active proteins and peptides, and several studies have described the characteristics of some of these toxins. However, complete proteomic profiling of the venoms of many snake species has not yet been done. The Indian cobra (Naja naja) and common krait (Bungarus caeruleus) are elapid snake species that are among the 'Big Four' responsible for the majority of human snake envenomation cases in India. As understanding the composition and complexity of venoms is necessary for successful treatment of envenomation in humans, we utilized three different proteomic profiling approaches to characterize these venoms: i) one-dimensional SDS-PAGE coupled with in-gel tryptic digestion and electrospray tandem mass spectrometry (ESI-LC-MS/MS) of individual protein bands; ii) in-solution tryptic digestion of crude venoms coupled with ESI-LC-MS/MS; and iii) separation by gel-filtration chromatography coupled with tryptic digestion and ESI-LC-MS/MS of separated fractions. From the generated data, 81 and 46 different proteins were identified from N. naja and B. caeruleus venoms, respectively, belonging to fifteen different protein families. Venoms from both species were found to contain a variety of phospholipases A2 and three-finger toxins, whereas relatively higher numbers of snake venom metalloproteinases were found in N. naja compared to B. caeruleus venom. The analyses also identified less represented venom proteins including L-amino acid oxidases, cysteine-rich secretory proteins, 5'-nucleotidases and venom nerve growth factors. Further, Kunitz-type serine protease inhibitors, cobra venom factors, phosphodiesterases, vespryns and aminopeptidases were identified in the N. naja venom, while acetylcholinesterases and hyaluronidases were found in the B. caeruleus venom. We further analyzed protein coverage (Lys/Arg rich and poor regions as well as potential glycosylation sites) using in-house software. These studies expand our understanding of the proteomes of the venoms of these two medically-important species.

On the Study of Pre-Pregnancy Body Mass Index (BMI) and Weight Gain as Indicators of Nutritional Status of Pregnant Women Belonging to Low Socio-Economic Category: A Study from Assam.

Women, particularly pregnant women, are the most vulnerable population of the society and their health status is one of the major indicators of development. There were enough studies on pre pregnancy body mass index (IPBMI) and inadequate weight gain during pregnancy (IWGP) of women in other part of the world and India, but none in Assam. In Assam a large number of population are in the category of low socio-economic group, a group most vulnerable to under nutrition. Thus this study was framed with the said indicators to throw light on the factors affecting the health status of pregnant women to accordingly address the situation. A cross sectional study using multistage sampling design with probability proportional to size was made comprising of 461 pregnant women belonging to low socio-economic status. Responses regarding their socio-economic, socio-cultural, health, diet and environmental background were collected and coded. The study revealed that although IPBMI (34.06%) was slightly lower than the reported state, national and global percentage the revealed IWGP (82%) was an astounding figure. The blood samples analyzed showed a high degree of inadequacy in almost all micronutrients (iron 63.1%, calcium 49.5% and copper 39.9%) studied in our survey.

Nutritional evaluation of popped and malted indigenous millet of Assam.

For better utilization of millets, two processing techniques, viz., popping and malting were standardized using two local varieties of foxtail millet (Setaria italica). In popped samples, crude fat and crude fibre contents were significantly lower than raw millet in both the yellow and purple varieties, while the carbohydrate and energy values were significantly higher. In malted samples, crude protein and fat contents were significantly lower than in raw millet in both the varieties, whereas the carbohydrate contents were higher. Starch digestibility was highest (42.4%) in yellow popped samples and lowest in yellow malted samples (21.8%). Protein digestibility was highest (13.2%) in purple popped and lowest (2.4%) in yellow malted samples.