Isoliquiritigenin: a potential drug candidate for the management of erectile dysfunction.

OBJECTIVE: This study aimed to assess the erectogenic properties of isoliquiritigenin taking sildenafil (SDF) as the standard. METHODS: The binding affinity of isoliquiritigenin (ISL) with the erectile marker proteins (endothelial nitric oxide synthase [eNOS] and enzyme phosphodiesterase type 5 [PDE5]) was investigated using Autodock Vina, which was validated using molecular dynamics simulation. Furthermore, the effect of ISL on the eNOS and PDE5 messenger ribonucleic acid (mRNA) expression and the sexual behavior of mice was investigated, along with the assessment of the pharmacokinetics of ISL. KEY FINDINGS: The results revealed that the binding affinity of ISL-eNOS/PDE5 and SDF-eNOS/PDE5 was in the range of -7.5 to -8.6 kcal/mol. The ISL-eNOS/PDE5 complexes remained stable throughout the 100 ns simulation period. Root mean square deviation, Rg, SASA, hydrogen, and hydrophobic interactions were similar between ISL-eNOS/PDE5 and SDF-eNOS/PDE5. Analysis of mRNA expressions in paroxetine (PRX)-induced ED mice showed that the co-administration of PRX with ISL reduced PDE5 and increased eNOS mRNA expression, similar to the co-administered group (PRX+SDF). The sexual behavior study revealed that the results of PRX+ISL were better than those of the PRX+SDF group. Pharmacokinetic evaluation further demonstrated that ISL possesses drug-like properties. CONCLUSIONS: The results showed that ISL is equally potent as SDF in terms of binding affinity, specific pharmacological properties, and modulating sexual behavior.

Scrub Typhus- An Underestimated Infectious Disease Attributable to Community Acquired Acute Kidney Injury.

Acute Kidney Injury (AKI) associated with Scrub typhus is an emerging health problem which is more common in the tropics including India. This study intended to find out the occurrence of Scrub typhus among the Community Acquired Acute Kidney Injury patients in a tertiary care hospital in Assam, North East India. AKI patients with acute febrile illness admitted to Gauhati Medical College and Hospital, Guwahati, Assam were included in the study and demographic characteristics along with clinical features were recorded. The detection of Scrub typhus was done by IgM Enzyme Linked Immunosorbent Assay (ELISA) test (Optical Density > 0.5) and polymerase chain reaction (PCR) analysis. Routine haematological and biochemical tests were performed. Molecular characterization of Orientia tsutsugamushi was done followed by phylogenetic analysis. The Graph Pad Prism software 9 was used for statistical analysis. Out of 221 AKI patients admitted to hospital, 45 patients (20.4%) were confirmed to be Scrub typhus positive and among them, 4 cases were co-infected with leptospirosis. Majority of Scrub typhus positive AKI patients were in Stage I (82.2%) under KDIGO guideline. "Karp" was the predominant circulating serotype. The study showed cases of Scrub typhus associated Acute Kidney Injury was high and mortality was 11.1%. Hence, in this region, further studies need to be done with large number of population and more emphasis need to be given on differential diagnosis. Supplementary Information: The online version contains supplementary material available at 10.1007/s12088-023-01137-x.

Co-circulation of two Mumps virus genotypes in Assam, India.

Mumps is a vaccine-preventable disease, and research on the vaccine's efficacy has recently indicated declining efficacy that has failed to protect against primary infections or reinfections, leading to a global resurgence in nations that use mumps vaccine in their national immunization programmes (NIPs). Lack of reports on its infection, documentation and published studies prevents it from being recognized as a public health issue in India. The waning of immunity is ascribed to the changes between the circulating and vaccine strains. The goal of the current study was to describe the circulating MuV strains in the Dibrugarh district of Assam, India, from 2016 to 2019. Blood samples were examined for IgM antibodies, and throat swab samples were put through Taqman assay for molecular detection. The small hydrophobic (SH) gene was targeted for genotyping through sequencing, and its genetic variations and phylogenetic analysis were carried out. Mumps RNA was found in 42 cases, and Mumps IgM in 14, of which 60% (25/42) of the cases were male and 40% (17/42) were female mostly affecting children between the ages of 6 and 12. Sequence and phylogeny analyses of SH gene revealed Genotypes C (83%) and G (17%) were simultaneously circulating during the study period. The study offers crucial genetic baseline information for the creation of Mumps prevention and control measures. Therefore, based on the research, it is clear that developing an effective vaccination strategy should take into account all currently prevalent genotypes in order to provide better protection against the disease's comeback.

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.

Exploring the anti-inflammatory potential of Colocasia esculenta root extract in in-vitro and in-vivo models of inflammation.

ETHNOPHARMACOLOGICAL RELEVANCE: Colocasia esculenta (CE) (L.) Schott is an annual herbaceous tropical plant from the family of Araceae which has been traditionally used for the healing of various ailments such as asthma, arthritis, internal hemorrhage, diarrhea, and neurological disorders. The plant is reported to have potential anti-microbial, anti-fungal, antimetastatic, anti-hepatotoxic, and anti-lipid peroxidative activities. AIM OF THE STUDY: The present study is designed to explore the potential anti-inflammatory property of Colocasia esculenta methanolic root extract (CEMRE) on carrageenan-induced rat paw edema and lipopolysaccharide (LPS) stimulated RAW264.7 cells. MATERIALS AND METHODS: Carrageenan-induced rat paw edema model was used to investigate the in vivo anti-inflammatory action of CEMRE. Adult male Wistar rats (180-220 g; n = 6) were pre-treated with CEMRE (100, 200, and 400 mg/kg BW) orally before 1 h of injection of 1% carrageenan. Indomethacin (10 mg/kg BW) was given orally as the standard drug. Serum aspartate aminotransferase (AST), alanine aminotransferase (ALT), alkaline phosphatase (ALP), nitric oxide (NO), prostaglandinE2 (PGE2), and cytokines levels were measured. Liquid chromatography-mass spectrometry (LC-MS) was done to identify the phytoconstituents present in CEMRE. The inhibitory activity of CEMRE was investigated against cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) in in vitro assessment of LPS-stimulated RAW264.7 cells. The RAW 264.7 cells were pre-treated with Indomethacin (5 µM and 10 µM) and CEMRE (17 µg/ml and 34 µg/ml) followed by induction of LPS (1 µg/ml) for 24 h. Docking analyses were also performed to explore the interaction of important phytoconstituents (Sinapic acid, Acetylsalicylic acid, L-fucose, Salicylic acid, Quinic acid, Zingerone, and Gingerol) of CEMRE with COX-2 and iNOS. RESULTS: Pre-treatment with CEMRE (400 mg/kg) could inhibit the paw inflammation significantly which was elevated due to carrageenan induction. The inhibition is comparable to that of the standard drug Indomethacin. The concentration of serum AST, ALT, ALP, NO, PGE2 and cytokines were also considerably lowered in the CEMRE-treated group as compared to the carrageenan-induced group. CEMRE (34 µg/ml) inhibited the LPS-stimulated relative expression of mRNA of COX-2 and iNOS and significantly reduced the expression of nitric oxide and prostaglandin E2. Docking analyses revealed promising interaction with low binding energies between Sinapic acid with both the target proteins COX-2 and iNOS. CONCLUSION: Collectively, our results suggested that CEMRE exhibited effective anti-inflammatory actions on carrageenan-induced rat paw edema and LPS-treated RAW 264.7 cells by reducing the in vivo paw edema inhibition, inhibiting the serum NO, PGE2, cytokines and also reduced the in vitro production of NO, PGE2 along with expressions of mRNA COX-2 and iNOS. Molecular docking demonstrated good binding affinities among the target proteins and ligand Sinapic acid. Thus the bioactive compound from CE need to be isolated and purified.

Genetic characterisation of influenza A(H1N1)pdm09 viruses circulating in Assam, Northeast India during 2009-2015.

Introduction: Influenza A(H1N1)pdm09 virus, since its identification in April 2009, has continued to cause significant outbreaks of respiratory tract infections including pandemics in humans. In the course of its evolution, the virus has acquired many mutations with an ability to cause increased disease severity. A regular molecular surveillance of the virus is essential to mark the evolutionary changes that may cause a shift to the viral behavior. Materials and Methods: Samples of Throat/Nasal swabs were collected from a total of 3715 influenza-like illness cases and screened by Real-time Reverse Transcription-Polymerase Chain Reaction for influenza viruses. Nucleotide sequence analysis was done to identify changes in antigenicity of the virus strains. Results: The present study describes the molecular characteristics of influenza A(H1N1)pdm09 viruses detected in Assam of Northeast India during 2009-2015. Influenza A viruses were detected in 11.4% (425/3715), of which influenza A(H1N1)pdm09 viruses were detected in 41.4% (176/425). The nucleotide sequencing of influenza A(H1N1)pdm09 viruses revealed a total of 17 and 22 amino acid substitutions in haemagglutinin (HA) and neuraminidase (NA) genes of the virus, respectively, compared to contemporary vaccine strain A/California/07/2009. The important mutations detected in HA genes of A/Assam(H1N1)pdm09 strains included E391K, K180Q and S202T. Mutation 'N248D' which has an ability to develop oseltamivir resistance was also detected in NA gene of A/Assam(H1N1)pdm09 strains. Conclusions: Regular molecular surveillance of influenza A(H1N1)pdm09 is important to monitor the viral behavior in terms of increase virulence, drug resistance pattern and emergence of novel strains.

Genetic variations of the Hemagglutinin gene of Pandemic Influenza A (H1N1) viruses in Assam, India during 2016.

Since its emergence in 2009, Influenza A/H1N1pdm09 virus has evolved continuously. Marked genetic variations have occurred in the HA1 domain of the hemagglutinin gene causing the emergence of new variants. The present study genetically characterized the hemagglutinin (HA) gene of Influenza A/H1N1pdm09 strains from Assam circulating in 2016 that caused a mild outbreak without any reported mortality. Sequence analysis of the HA gene of 20 positive Assam/H1N1pdm09 strains revealed 3 mutations (K180Q, S202T, S220T) at the antigenic sites along with several other reported mutations which are in close proximity to the antigenic sites and therefore might affect the viral antigenicity. Phylogenetically, the Assam/H1N1pdm09 strains clustered into genogroup 6B. These genetic variations highlight the importance of continuous surveillance and characterization of Influenza A/H1N1pdm09 virus activity to track the genetic makeup and diversification that may affect the behavior of the virus.

Molecular characterisation and phylogenetic analysis of dengue outbreak in Pasighat, Arunachal Pradesh, Northeast India.

BACKGROUND AND OBJECTIVES: Dengue is one of the most prevalent arboviral diseases in the world with 390 million dengue infections per year. In this study, we report the molecular characterisation of dengue outbreak in Pasighat, Arunachal Pradesh, Northeast India during 2015. SUBJECTS AND METHODS: : A total of 613 dengue-suspected cases were screened for dengue virus by dengue NS1 Ag and anti-dengue IgM antibody depending on the duration of sample collection and onset of symptom. Further, molecular characterisation was done by amplifying the C-PrM region by real-time polymerase chain reaction followed by phylogenetic analysis. RESULTS: Molecular characterisation revealed that the dengue outbreak was predominantly due to dengue virus serotype-1 (DENV-1) (90.9%) while DENV-2 was detected in 7.5% of samples. Co-infection of DENV-1 and DENV-2 was detected in one case. Phylogenetic analysis of the DENV-1 strains with the prototype revealed that the DENV-1 strains were grouped within genotype III. Similarly, DENV-2 strains were clustered within genotype IV. The study revealed a change in the predominant serotype in recent years with DENV-3 in 2012 to DENV-1, 2, 3 and 4 in 2014 to DENV-1 in 2015 in the study region. A unique L24M mutation was observed in the DENV-1 strains of Arunachal Pradesh which was absent in all the circulating strains in India except one strain from the state of Kerala in South India. Marked variation within the DENV-2 strains was observed at A102V and I163V in one strain similar to earlier circulating isolates in India. CONCLUSIONS: The present study reveals a shift in the serotype dominance in the study region. As serotype shifts and secondary infection with a heterologous DENV serotype are frequently associated with disease severity, there is an urgent need for sustained monitoring of the circulating serotypes and enhanced surveillance operations, especially in the monsoon and post-monsoon periods to prevent large-scale, severe dengue outbreaks in this region.

Design, in silico and in vitro evaluation of curcumin analogues against Plasmodium falciparum.

The polyphenolic compound curcumin has been reported for its antimalarial properties in various scientific studies. Plasmodium falciparum ATP6, the parasite orthologue of mammalian sarcoplasmic Ca2+ ATPase (SERCA) has been identified as a key molecular target of both artemisinin and curcumin. The work was thereby undertaken to study the anti-malarial properties of two different series of curcumin analogues based on their docking interactions with PfATP6 and correlating the results with their anti-malarial activity. The compounds were designed retaining similar functional groups as that of the parent curcumin nucleus while incorporating changes in the carbon chain length, unsaturated groups and the number of ketone groups. The compounds (1E, 4E)-1,5-bis(4-methylphenyl)penta-1,4-dien-3-one (CD-9), (1E, 4E)-1,5-bis(4-methoxyphenyl)penta-1,4-dien-3-one (CD-8) and (E)-1,3-bis(4-hydroxylphenyl)prop-2-en-1-one (CD-1) showed IC50 values of 1.642 µM, 1.764 µM and 2.59 µM in 3D7 strain and 3.039 µM, 7.40 µM and 11.3 µM in RKL-2 strain respectively. Detailed structure-activity relationship studies of the compounds showed that CD-9 and CD-8 had a common hydrophobic interaction with the residue Leu268 of the PfATP6 protein and has been postulated through our study to be the reason for their antimalarial activity as seen after corroborating the results with the in vitro study. The study provided valuable insight about the ligand-protein interaction of the various functional groups of curcumin and its analogues against the PfATP6 protein and their importance in imparting antimalarial action.

Molecular Modeling and Dynamics Simulation Analysis of KATNAL1 for Identification of Novel Inhibitor of Sperm Maturation.

BACKGROUND: Hormone based birth control often causes various side effects. A recent study revealed that temporary infertility without changing hormone levels can be attained by inhibiting Katanin p60 ATPase-containing subunit A-like 1 protein (KATNAL1) which is critical for sperm maturation in the testes. OBJECTIVE: This study aimed at attaining the most energetically stable three dimensional (3D) structure of KATNAL1 protein using comparative modeling followed by screening of a ligand library of known natural spermicidal compounds for their binding affinity with KATNAL1. This in turn may inhibit the development of mature sperm in the seminiferous epithelium. METHOD: A series of computational techniques were used for building the 3D structure of KATNAL1 which was further optimized by molecular dynamics (MD) simulation. For revealing the ATP binding mode of KATNAL1, docking study was carried out using the optimized model obtained from the MD simulation. The docking study was also employed to test the binding efficiency of the ligand library. RESULTS: Molecular docking study confirmed the ATP binding of KATNAL1 with various hydrophobic and hydrogen bond interactions. Binding efficiency of the ligand library suggested that calotropin, a cardenolide of Calotropis procera showed the highest binding efficiency against the target protein without toxicity. MD simulation of the docked complex validated the results of the docking study. CONCLUSION: This study revealed the ATP binding mode of KATNAL1 and identified calotropin as a potential lead molecule against it showing high binding efficiency with good bioavailability and no mutagenicity. Further in vitro and in vivo bioassay of calotropin could facilitate the development of novel non-hormonal male-specific contraceptive in near future.

Molecular characterization of Plasmodium falciparum in Arunachal Pradesh from Northeast India based on merozoite surface protein 1 & glutamate-rich protein.

BACKGROUND & OBJECTIVES: Northeast (NE) India is one of the high endemic regions for malaria with a preponderance of Plasmodium falciparum, resulting in high morbidity and mortality. The P. falciparum parasite of this region showed high polymorphism in drug-resistant molecular biomarkers. However, there is a paucity of information related to merozoite surface protein 1 (msp-1) and glutamate-rich protein (glurp) which have been extensively studied in various parts of the world. The present study was, therefore, aimed at investigating the genetic diversity of P. falciparum based on msp-1 and glurp in Arunachal Pradesh, a State in NE India. METHODS: Two hundred and forty nine patients with fever were screened for malaria, of whom 75 were positive for P. falciparum. Blood samples were collected from each microscopically confirmed patient. The DNA was extracted; nested polymerase chain reaction and sequencing were performed to study the genetic diversity of msp-1 (block 2) and glurp. RESULTS: The block 2 of msp-1 gene was found to be highly polymorphic, and overall allelic distribution showed that RO33 was the dominant allele (63%), followed by MAD20 (29%) and K1 (8%) alleles. However, an extensive diversity (9 alleles and 4 genotypes) and 6-10 repeat regions exclusively of R2 type were observed in glurp. INTERPRETATION & CONCLUSIONS: The P. falciparum population of NE India was diverse which might be responsible for higher plasticity leading to the survival of the parasite and in turn to the higher endemicity of falciparum malaria of this region.

Molecular docking, synthesis and in vitro antimalarial evaluation of certain novel curcumin analogues

ABSTRACT The receptor protein PfATP6 has been identified as the common target of artemisinin and curcumin. The work was initiated to assess the antimalarial activity of six curcumin derivatives based on their binding affinities and correlating the in silico docking outcome with in vitro antimalarial screening results. A ligand library of thirty two Knoevenagel condensates of curcumin were designed and docked against PfATP6 protein and six compounds with the best binding scores were synthesized and screened for their antimalarial activity against the sensitive 3D7 strain of Plasmodium falciparum. ADME/Tox, pharmacokinetic and pharmacodynamic profiles of the designed compounds were analyzed and reported. 4-FB was found to have similar binding energy to the standard artemisinin (-6.75 and -6.73 respectively) while 4-MB, 3-HB, 2-HB, B, 4-NB displayed better binding energy than curcumin (-5.95, -5.89, -5.68, -5.35, -5.29 and -5.25 respectively). At a dose of 50 µg/mL all the six compounds showed 100% schizont inhibition while at 5µg/ml, five showed more than 75% inhibition and better results than curcumin. 4-FB showed the best activity with 97.8% schizonticidal activity. The in vitro results superimpose the results obtained from the in silico study thereby encouraging development of promising curcumin leads in the battle against malaria.

Metagenomics analysis of microbial communities associated with a traditional rice wine starter culture (Xaj-pitha) of Assam, India.

This is the first report on the microbial diversity of xaj-pitha, a rice wine fermentation starter culture through a metagenomics approach involving Illumine-based whole genome shotgun (WGS) sequencing method. Metagenomic DNA was extracted from rice wine starter culture concocted by Ahom community of Assam and analyzed using a MiSeq® System. A total of 2,78,231 contigs, with an average read length of 640.13 bp, were obtained. Data obtained from the use of several taxonomic profiling tools were compared with previously reported microbial diversity studies through the culture-dependent and culture-independent method. The microbial community revealed the existence of amylase producers, such as Rhizopus delemar, Mucor circinelloides, and Aspergillus sp. Ethanol producers viz., Meyerozyma guilliermondii, Wickerhamomyces ciferrii, Saccharomyces cerevisiae, Candida glabrata, Debaryomyces hansenii, Ogataea parapolymorpha, and Dekkera bruxellensis, were found associated with the starter culture along with a diverse range of opportunistic contaminants. The bacterial microflora was dominated by lactic acid bacteria (LAB). The most frequent occurring LAB was Lactobacillus plantarum, Lactobacillus brevis, Leuconostoc lactis, Weissella cibaria, Lactococcus lactis, Weissella para mesenteroides, Leuconostoc pseudomesenteroides, etc. Our study provided a comprehensive picture of microbial diversity associated with rice wine fermentation starter and indicated the superiority of metagenomic sequencing over previously used techniques.

Structural comparison, substrate specificity, and inhibitor binding of AGPase small subunit from monocot and dicot: present insight and future potential.

ADP-glucose pyrophosphorylase (AGPase) is the first rate limiting enzyme of starch biosynthesis pathway and has been exploited as the target for greater starch yield in several plants. The structure-function analysis and substrate binding specificity of AGPase have provided enormous potential for understanding the role of specific amino acid or motifs responsible for allosteric regulation and catalytic mechanisms, which facilitate the engineering of AGPases. We report the three-dimensional structure, substrate, and inhibitor binding specificity of AGPase small subunit from different monocot and dicot crop plants. Both monocot and dicot subunits were found to exploit similar interactions with the substrate and inhibitor molecule as in the case of their closest homologue potato tuber AGPase small subunit. Comparative sequence and structural analysis followed by molecular docking and electrostatic surface potential analysis reveal that rearrangements of secondary structure elements, substrate, and inhibitor binding residues are strongly conserved and follow common folding pattern and orientation within monocot and dicot displaying a similar mode of allosteric regulation and catalytic mechanism. The results from this study along with site-directed mutagenesis complemented by molecular dynamics simulation will shed more light on increasing the starch content of crop plants to ensure the food security worldwide.

In silico screening of antifolate based novel inhibitors from Brucea mollis Wall. ex kurz against quadruple mutant drug resistant PfDHFR.

Plasmodium falciparum is the most lethal form of the genus Plasmodium which causes malaria, a 'disease of antiquity'. Globally it affects the health and socio-economic development of a large population especially in Sub-Saharan Africa and Southeast Asia. The Plasmodium falciparum dihydrofolate reductase-thymidylate synthase (PfDHFR-TS) is an important target of antimalarial drugs. Mutations at the active site of PfDHFR have resulted in decrease drug binding affinity of DHFR-inhibitors. In the present study we selected ten compounds of Brucea mollis Wall. Ex kurz and checked for their drug likeness using various computational tools and potential interactions with PfDHFR by molecular docking study. Soulameanone, a quassinoid of Brucea mollis Wall. Ex kurz showed better binding affinity when compared to pyrimethamine for both wild and quadruple mutant drug resistant PfDHFR. In addition, similar isomers of soulameanone were screened for their drug likeness and to study their interactions with PfDHFR. Twenty three compounds showed better binding affinity compared to soulameanone.

Insight into structural organization and protein-protein interaction of non structural 3 (NS3) proteins from dengue serotypes.

Dengue infections produce a distinct character of virus-induced intracellular membrane alterations which are associated with the viral replication machinery. Currently, the NS3 protein is being targeted for antiviral therapy against dengue. NS3 protein of dengue virus interacts with nuclear receptor binding protein (NRBP) of human causing cell trafficking between the Endoplasmic Reticulum (ER) and Golgi, which interacts with Rac3, a member of the Rho-GTPase family. No crystal structure of the NRBP is available for any species, thus limiting the complete understanding of structure- function relationships of this protein. The present study deals with the molecular modeling of the viral protein (NS3 of DENV1-4), the host protein (NRBP) and their interactions through protein-protein docking study. Theoretical threedimensional structures of the NRBP and NS3 were modeled using the Modeller 9v8, and the evaluated models were docked using GRAMM-X to study the mode of protein-protein interaction (NRBP as receptor and NS3 as ligand). The docked docking complexes were further evaluated for interaction analysis by the RosettaDock Server. Suface and interface residues were observed along with hydrogen and hydrophobic interaction. The conserved residues forming hydrogen interaction of NRBP with DENV1-4 serotypes were found to be GLN 305, SER 363 and GLN 379.

Design, virtual screening and docking study of novel NS3 inhibitors by targeting protein-protein interacting sites of dengue virus - a novel approach.

Currently dengue is a serious disease which has become a global burden in the last decade. Unfortunately, there are no effective drugs and vaccines against this disease. DENV non-structural protein (NS) 3, which is viral protease which is a potential target for antiviral therapy. Targeting this we performed homology modeling and protein-protein docking study of NS3 with NRBP (Nuclear Receptor Binding Protein) of human as it has been proved that NS3 of DENV interacts with NRBP which causes cellular trafficking in human cell. To carry out search of novel DENV protease inhibitors by in silico screening panduratin molecule was selected. 65 novel compounds were designed which involved substituting positions 1-5 of the benzyl ring A (4hydroxy-panduratinA) with various substituents. The protein-protein docking showed that the aminoacid residues of NS3 which were interacting with NRBP were found to be Ala 325, Asp 324, Phe 326, Asp 335, Glu 336, Glu 328, Asp 485, Gln 478, Arg 459, Gly 446 and Leu 480. These residues were targeted by the ligands which showed excellent binding affinity as binding energy. The ligand PKP10 showed lowest binding energy. It is also observed that the interface residues participated in the protein-protein interaction are being inhibited by the ligands.

Structural analysis and molecular dynamics simulations of novel δ-endotoxin Cry1Id from Bacillus thuringiensis to pave the way for development of novel fusion proteins against insect pests of crops.

The theoretical three-dimensional structure of a novel δ-endotoxin Cry1Id (81 kDa) belonging to Cry1I class, toxic to many of the lepidopteran pests has been investigated through comparative modeling. Molecular dynamics (MD) simulations was carried out to characterize its structural and dynamical features at 10 ns in explicit solvent using the GROMACS version 4.5.4. Finally the simulated model was validated by the SAVES, WHAT IF, MetaMQAP, ProQ, ModFOLD and MolProbity servers. Despite low sequence identity with its structural homologs, Cry1Id not only resembles the previously reported Cry structures but also shares the common five conserved blocks of amino acid residues. Although the domain II of Cry1Id superpose well with its closest structural homolog Cry8Ea1, variation of amino acids and length in the apical loop2 of domain II was observed. In this work, we have hypothesized that the variations in apical loop2 might be the sole factor for providing variable surface accessibility to Cry1Id protein that could be important in receptor recognition. MD simulation showed the proposed endotoxin retains its stable conformation in aqueous solution. The result from this study is expected to aid in the development hybrid Cry proteins and new potent fusion proteins with novel specificities against different insect pests for improved pest management of crop plants.

Insights into the structure-function relationship of disease resistance protein HCTR in maize (Zea mays L.): a computational structural biology approach.

The disease resistance gene Hm1 of maize encodes a NADPH-dependent reductase enzyme, HC-toxin reductase (HCTR) that detoxifies the HC toxin secreted by the race specific fungus Cochliobolus carbonum race 1. HCTR enzyme shares 29.6% sequence identity with dihydroflavonol reductase (DFR) of grape, a key enzyme involved in flavonoid biosynthesis. Here we report the comparative modelling, molecular dynamics simulation and docking studies to explain the structure-function relationship and the mode of cofactor (NADPH) binding in HCTR enzyme at the molecular level. The nucleotide binding domain of modelled HCTR adopts a classic Rossmann fold and possesses a consensus glycine rich GxGxxG motif. Molecular simulation studies suggested that HCTR model retained stability throughout the simulation in aqueous solution. HCTR model showed considerable structural identities with the cofactor binding site of DFR, but significant difference in the catalytic site might be the reason of functional divergence between these families of proteins. Similarly electrostatic surface potential analysis of both HCTR and DFR revealed profound variations in the charge distribution over the substrate binding site, which can be correlated with the sequence variability and may suggest distinct substrate-binding patterns and differences in the catalytic mechanism. Docking results indicated Phe19, Gly21, Arg40, Thr90, Gly208, Arg218, Glu221 and Thr222 are important residues for cofactor (NADPH) binding through strong hydrogen bonding and electrostatic interactions. Alanine scanning and analysis of docking energies of mutant proteins suggested that Phe19, and Arg40 are two critical residues for the cofactor binding. The result from the present study is expected to pave the way for exploration of similar genes in other economically important crop varieties.

In silico identification and characterization of conserved miRNAs and their target genes in sweet potato (Ipomoea batatas L.) expressed sequence tags (ESTs).

The endogenous small non-coding micro RNAs (miRNAs), which are typically ~21-24 nt nucleotides, play a crucial role in regulating the intrinsic normal growth of cells and development of the plants as well as in maintaining the integrity of genomes. These small non-coding RNAs function as the universal specificity factors in post-transcriptional gene silencing. Discovering miRNAs, identifying their targets, and further inferring miRNA functions is a routine process to understand normal biological processes of miRNAs and their roles in the development of plants. Comparative genomics based approach using expressed sequence tags (EST) and genome survey sequences (GSS) offer a cost-effective platform for identification and characterization of miRNAs and their target genes in plants. Despite the fact that sweet potato (Ipomoea batatas L.) is an important staple food source for poor small farmers throughout the world, the role of miRNA in various developmental processes remains largely unknown. In this paper, we report the computational identification of miRNAs and their target genes in sweet potato from their ESTs. Using comparative genomics-based approach, 8 potential miRNA candidates belonging to miR168, miR2911, and miR156 families were identified from 23 406 ESTs in sweet potato. A total of 42 target genes were predicted and their probable functions were illustrated. Most of the newly identified miRNAs target transcription factors as well as genes involved in plant growth and development, signal transduction, metabolism, defense, and stress response. The identification of miRNAs and their targets is expected to accelerate the pace of miRNA discovery, leading to an improved understanding of the role of miRNA in development and physiology of sweet potato, as well as stress response.