Identification of potential inhibitor against Leishmania donovani mitochondrial DNA primase through in-silico and in vitro drug repurposing approaches.

Leishmania donovani is the causal organism of leishmaniasis with critical health implications affecting about 12 million people around the globe. Due to less efficacy, adverse side effects, and resistance, the available therapeutic molecules fail to control leishmaniasis. The mitochondrial primase of Leishmania donovani (LdmtPRI1) is a vital cog in the DNA replication mechanism, as the enzyme initiates the replication of the mitochondrial genome of Leishmania donovani. Hence, we target this protein as a probable drug target against leishmaniasis. The de-novo approach enabled computational prediction of the three-dimensional structure of LdmtPRI1, and its active sites were identified. Ligands from commercially available drug compounds were selected and docked against LdmtPRI1. The compounds were chosen for pharmacokinetic study and molecular dynamics simulation based on their binding energies and protein interactions. The LdmtPRI1 gene was cloned, overexpressed, and purified, and a primase activity assay was performed. The selected compounds were verified experimentally by the parasite and primase inhibition assay. Capecitabine was observed to be effective against the promastigote form of Leishmania donovani, as well as inhibiting primase activity. This study's findings suggest capecitabine might be a potential anti-leishmanial drug candidate after adequate further studies.

Isolation, identification, and molecular characterization of potential keratinolytic fungus sp. from Southern Assam: relevance to poultry wastes and its biological management.

Feather waste is a highly prevalent form of keratinous waste that is generated by the poultry industry. The global daily production of feather waste has been shown to approach 5 million tons, typically being disposed of through methods such as dumping, landfilling, or incineration which contribute significantly to environmental pollutions. The proper management of these keratinous wastes is crucial to avoid environmental contamination. The study was carried out to isolate the keratinolytic fungi from the poultry disposal sites of different region of North-East India to evaluate its potential in bioremediation of the feathers wastes. Out of 12 fungal strains isolated from the sites, the fungus showing the highest zone of hydrolysis on both the skim milk and keratin agar medium was selected for the study and the molecular identification of the isolate was performed through DNA sequence analysis by amplifying the internal transcribed spacer (ITS) region. The sequence results showed higher similarity (above 95%) with Aspergillus spp. and was named Aspergillus sp. Iro-1. The strain was further analyzed for its feather degrading potential which was performed in submerged conditions under optimized conditions. The study showed that the strain could effectively degrade the feathers validated through weight loss method, and the structural deformations in the feathers were visualized through scanning electron microscopy (SEM). Aspergillus sp. Iro-1 was obtained from the southern region of Assam. It would be of great importance as the implementation of this sp. can help in the bioremediation of feathers wastes in this region. This is the first study of identification of feather degrading fungus from southern part of Assam (Barak).

Green synthesis of novel biochar from Abelmoschus esculentus seeds for direct blue 86 dye removal: Characterization, RSM optimization, isotherms, kinetics, and fixed bed column studies.

The presence of Direct blue 86 dye (DB86) in water can lead to various health hazards to the humans and animals. The study explored efficacy of biochar derived from Abelmoschus Esculentus seeds (AESB) to remove DB86 from an aqueous solution. BET analysis of AESB delineated H4 classification with the predominance of micropores and mesopores spread throughout the surface. FTIR study demonstrated the presence of the alkyl (C-H), Alkene (C]C), Carbonyl (C]O) and O-H bond of the sulphonic group which helped in adsorption of DB86 molecules through various mechanisms i.e., pore filling, π-π interactions, and hydrogen bonding interactions. Response surface methodology (RSM) was used for designing the adsorption experiment and analysing the optimum operating parameters. Batch experiments demonstrated excellent adsorption capacity (277.04 mg/g) of AESB and was efficient in 98.06% removal of DB86 at optimal conditions i.e., dye conc. = 300 mg/L, dose = 2.5 g/L, pH = 2, time of 120 min. Adsorption followed nonlinear Sips model (R2 = 0.999) with an error (X2 = 0.13, RMSE = 0.83, MAPE 0.56 and MSRE = 0.0006). The kinetic analysis revealed intra-particle diffusion being the rate-determining step and followed nonlinear pseudo-first-order kinetics (R2 = 0.997). Thermodynamic study revealed that the adsorption of DB-86 proceeded spontaneously and exhibited endothermic characteristics, with the enthalpy change primarily governed by the physisorption mechanism. Thomas model revealed inverse relation of breakthrough and exhaustion time with flow while it was proportional to bed height. The sorption capacity (N0) (2.2493 mg/l min) and rate constant (Ka) (0.028 L/min. mg) of BDST model can accurately be used for predicting the performance of AESB in full scale column.

In silico and immunoinformatics based multiepitope subunit vaccine design for protection against visceral leishmaniasis.

Visceral leishmaniasis (VL) is a vector-borne neglected tropical protozoan disease with high fatality and no certified vaccine. Conventional vaccine preparation is challenging and tedious. Here in this work, we created a global multiepitope subunit vaccination against VL utilizing innovative immunoinformatics technique based on the extensively conserved epitopic regions of the PrimPol protein of Leishmania donovani consisting of four subunits which were analyzed and studied, out of which DNA primase large subunit and DNA polymerase α subunit B were evaluated as antigens by Vaxijen 2.0. The multiepitope vaccine design includes a single adjuvant ß-defensins, eight CTL epitopes, eight HTL epitopes, seven linear BCL epitopes and one discontinuous BCL epitope to induce innate, cellular and humoral immune responses against VL. The Expasy ProtParam tool characterized the physiochemical parameters of the vaccine. At the same time, SOLpro evaluated our vaccine constructs to be soluble upon expression. We also modeled the stable tertiary structure of our vaccine construct through Robetta modeling for molecular docking studies with toll-like receptor proteins through HADDOCK 2.4. Simulations based on molecular dynamics revealed an intact vaccine and TLR8 complex, supporting our vaccine design's immunogenicity. Also, the immune simulation of our vaccine by the C-ImmSim server demonstrated the potency of the multiepitope vaccine construct to induce proper immune response for host defense. Codon optimization and in silico cloning of our vaccine further assured high expression. The outcomes of our study on multiepitope vaccine design significantly produced a potential candidate against VL and can potentially eradicate the disease in the future after clinical investigations.Communicated by Ramaswamy H. Sarma.

Synthesis and biological evaluation of carbohydrate-Coumarin/vanillic acid hybrid as a promising antiparasitic agent.

Leishmaniasis is caused by infection with the protozoan parasites Leishmania. It is classified as one of the most significant neglected tropical diseases. It remains a significant global public health concern. Current treatments include the use of pentavalent antimonial, amphotericin B, pentamidine, miltefosine, and paromomycin. However, several limitations such as toxicity, side effect, and resistance to these drugs of certain species are of concern. To combat this disease, effective chemotherapy is urgently required for its treatment and management. In this study, we synthesized a series of carbohydrate-coumarin/vanillic acid hybrids linked through triazole moiety via CuACC (Copper-catalysed azide-alkyne cycloaddition) reaction. These compounds were evaluated for their in vitro antiparasitic activity using MTT assay against Leishmania donovani whereas, all compounds show IC50 value in the range of 65-74 µM.

Drug repurposing based novel anti-leishmanial drug screening using in-silico and in-vitro approaches.

Visceral leishmaniasis is a neglected tropical disease and is mainly caused by L. donovani in the Indian subcontinent. The mitochondria genome replication in Leishmania spp. is having a very specific mechanism, and it is initiated by a key enzyme called mitochondrial primase. This enzyme is essential for the onset of the replication process and growth of the parasite. Therefore, we focused on the primase protein as a potential therapeutic target for combating leishmaniasis diseases. We started our studies molecular modeling and followed by docking of the FDA-approved drug library into the binding site of the primase protein. The top 30 selected compounds were subjected for molecular dynamics studies. Also, the target protein was cloned, purified, and tested experimentally (primase activity assays and inhibition assays). Some compounds were very effective against the Leishmania cell culture. All these approaches helped us to identify few possible novel anti-leishmanial drugs such as Pioglitazone and Mupirocin. These drugs are effectively involved in inhibiting the promastigote of L. donovani, and it can be utilized in the next level of clinical trials. Communicated by Ramaswamy H. Sarma.

Synthesis and inhibition studies towards the discovery of benzodiazepines as potential antimalarial compounds.

The target-based discovery of therapeutics against apicoplast, an all-important organelle is an overriding perspective. MEP pathway, an accredited drug target provides an insight into the importance of apicoplast in the survival of the parasite. In this study, we present the rational design strategy employing sustainable catalysis for the synthesis of benzodiazepine (BDZ) conformers followed by their biological evaluation as prospective inhibitors against the potential target of the IPP pathway, 1-deoxy-D-xylulose-5-phosphatereductoisomerase (DXR). The study reported the inhibitory profile of 8c and 6d against the quintessential step of the only drug target in the erythrocytic stages of parasite development. The potential compounds were identified to represent a novel class of inhibitors that serve as the lead molecules to impede the pathway and further affect the survival of the parasite.

Garcinia morella extract confers dopaminergic neuroprotection by mitigating mitochondrial dysfunctions and inflammation in mouse model of Parkinson's disease.

Dopaminergic neuroprotection is the main interest in designing novel therapeutics against Parkinson's disease (PD). In the process of dopaminergic degeneration, mitochondrial dysfunctions and inflammation are significant. While the existing drugs provide symptomatic relief against PD, a therapy conferring total neuroprotection by targeting multiple degenerative pathways is still lacking. Garcinia morella is a common constituent of Ayurvedic medication and has been used for the treatment of inflammatory disorders. The present study investigates whether administration of G. morella fruit extract (GME) in MPTP mouse model of PD protects against dopaminergic neurodegeneration, including the underlying pathophysiologies, and reverses the motor behavioural abnormalities. Administration of GME prevented the loss of dopaminergic cell bodies in the substantia nigra and its terminals in the corpus striatum of PD mice. Subsequently, reversal of parkinsonian behavioural abnormalities, viz. akinesia, catalepsy, and rearing, was observed along with the recovery of striatal dopamine and its metabolites in the experimental model. Furthermore, reduced activity of the mitochondrial complex II in the nigrostriatal pathway of brain of the mice was restored after the administration of GME. Also, MPTP-induced enhanced activation of Glial fibrillary acidic protein (GFAP) and neuronal nitric oxide synthase (nNOS) in the nigrostriatal pathway, which are the markers of inflammatory stress, were found to be ameliorated on GME treatment. Thus, our study presented a novel mode of dopaminergic neuroprotection by G. morella in PD by targeting the mitochondrial dysfunctions and neuroinflammation, which are considered to be intricately associated with the loss of dopaminergic neurons.

Pharmacogenomic landscape of Indian population using whole genomes.

Ethnic differences in pharmacogenomic (PGx) variants have been well documented in literature and could significantly impact variability in response and adverse events to therapeutics. India is a large country with diverse ethnic populations of distinct genetic architecture. India's national genome sequencing initiative (IndiGen) provides a unique opportunity to explore the landscape of PGx variants using population-scale whole genome sequences. We have analyzed the IndiGen variation dataset (N = 1029 genomes) along with global population scale databases to map the most prevalent clinically actionable and potentially deleterious PGx variants among Indians. Differential frequencies for the known and novel variants were studied and interaction of the disrupted PGx genes affecting drug responses were analyzed by performing a pathway analysis. We have highlighted significant differences in the allele frequencies of clinically actionable PGx variants in Indians when compared to the global populations. We identified 134 mostly common (allele frequency [AF] > 0.1) potentially deleterious PGx variants that could alter or inhibit the function of 102 pharmacogenes in Indians. We also estimate that on, an average, each Indian individual carried eight PGx variants (single nucleotide variants) that have a direct impact on the choice of treatment or drug dosing. We have also highlighted clinically actionable PGx variants and genes for which preemptive genotyping is most recommended for the Indian population. The study has put forward the most comprehensive PGx landscape of the Indian population from whole genomes that could enable optimized drug selection and genotype-guided prescriptions for improved therapeutic outcomes and minimizing adverse events.

In-silico studies on Myo inositol-1-phosphate synthase of Leishmania donovani in search of anti-leishmaniasis.

Myo-inositol is one of the vital nutritional requirements for the Leishmania parasites' survival and virulence in the mammalian host. . Myo-inositol-1-phosphate synthase (MIPS) is responsible for the synthesis of myo-inositol in Leishmania, which plays a vital role in Leishmania's virulence to mammalian hosts. Earlier studies suggest MIP synthase as a potential drug target against which valproate was used as a drug. So, MIP synthase can be used as a target for anti-leishmanial drugs, and its inhibition may help in preventing leishmaniasis. The present study aims to identify valproate's potent analogs as drugs against MIP synthase of L. donovani (Ld-MIPS) with minimum side effects and toxicity to host.In this study, the three-dimensional structure of Ld-MIPS was built, followed by active site prediction. Ligand-based virtual screening was done using hybrid similarity recognition methods. The best 123 valproate analogs were filtered based on their quantitative structure activity relationship (QSAR) properties and were docked against Ld-MIPS using FlexX, PyRx and iGEMDOCK software. The topmost five ligands were selected for molecular dynamics simulation and pharmacokinetic analysis based on the docking score. Simulation studies up to 30 ns revealed that all five lead molecules bound with Ld-MIPS throughout MD simulation and there was no variation in their backbone. All the chosen inhibitors exhibited good pharmacokinetics/ADMET predictions with an excellent absorption profile, metabolism, oral bioavailability, solubility, excretion, and minimal toxicity, suggesting that these inhibitors may further be developed as anti-leishmaniasis drugs to prevent the spread of leishmaniasis.Communicated by Ramaswamy H. Sarma.

Discovering potential inhibitors against SARS-CoV-2 by targeting Nsp13 Helicase.

The rise in the incidence of COVID-19 as a result of SARS-CoV-2 infection has threatened public health globally. Till now, there have been no proper prophylactics available to fight COVID-19, necessitating the advancement and evolution of effective curative against SARS-CoV-2. This study aimed at the nonstructural protein 13 (nsp13) helicase as a promising target for drug development against COVID-19. A unique collection of nucleoside analogs was screened against the SARS-CoV-2 helicase protein, for which a molecular docking experiment was executed to depict the selected ligand's binding affinity with the SARS-CoV-2 helicase proteins. Simultaneously, molecular dynamic simulations were performed to examine the protein's binding site's conformational stability, flexibility, and interaction with the ligands. Key nucleoside ligands were selected for pharmacokinetic analysis based on their docking scores. Selected ligands (cordycepin and pritelivir) showed excellent pharmacokinetics and were well stabilized at the proteins' binding site throughout the MD simulation. We have also performed binding free energy analysis or the binding characteristics of ligands with Nsp13 by using MM-PBSA and MM-GBSA. Free energy calculation by MM-PBSA and MM-GBSA analysis suggests that pritelivir may work as viable therapeutics for efficient drug advancement against SARS-CoV-2 Nsp13 helicase, potentially arresting the SARS-CoV-2 replication.Communicated by Ramaswamy H. Sarma.

Discovering Potential RNA Dependent RNA Polymerase Inhibitors as Prospective Drugs Against COVID-19: An in silico Approach.

COVID-19, caused by Severe Acute Respiratory Syndrome Corona Virus 2, is declared a Global Pandemic by WHO in early 2020. In the present situation, though more than 180 vaccine candidates with some already approved for emergency use, are currently in development against SARS-CoV-2, their safety and efficacy data is still in a very preliminary stage to recognize them as a new treatment, which demands an utmost emergency for the development of an alternative anti-COVID-19 drug sine qua non for a COVID-19 free world. Since RNA-dependent RNA polymerase (RdRp) is an essential protein involved in replicating the virus, it can be held as a potential drug target. We were keen to explore the plant-based product against RdRp and analyze its inhibitory potential to treat COVID-19. A unique collection of 248 plant compounds were selected based on their antiviral activity published in previous literature and were subjected to molecular docking analysis against the catalytic sub-unit of RdRp. The docking study was followed by a pharmacokinetics analysis and molecular dynamics simulation study of the selected best-docked compounds. Tellimagrandin I, SaikosaponinB2, Hesperidin and (-)-Epigallocatechin Gallate were the most prominent ones that showed strong binding affinity toward RdRp. All the compounds mentioned showed satisfactory pharmacokinetics properties and remained stabilized at their respective binding sites during the Molecular dynamics simulation. Additionally, we calculated the free-binding energy/the binding properties of RdRp-ligand complexes with the connection of MM/GBSA. Interestingly, we observe that SaikosaponinB2 gives the best binding affinity (∆Gbinding = -42.43 kcal/mol) in the MM/GBSA assay. Whereas, least activity is observed for Hesperidin (∆Gbinding = -22.72 kcal/mol). Overall our study unveiled the feasibility of the SaikosaponinB2 to serve as potential molecules for developing an effective therapy against COVID-19 by inhibiting one of its most crucial replication proteins, RdRp.

Evaluation of flavonoids as 2019-nCoV cell entry inhibitor through molecular docking and pharmacological analysis.

The outbreak of Coronavirus Disease 2019 (COVID-19) has been declared as a Public Health Emergency of International Concern (PHEIC) by the World Health Organization (WHO), which is being rapidly spread by the extremely spreadable and pathogenic 2019 novel coronavirus (2019-nCoV), also known as SARS-CoV-2. Pandemic incidence of COVID-19 has created a severe threat to global public health, necessitating the development of effective drugs or inhibitors or therapeutics agents against SARS-CoV-2. Spike protein (S) of the SARS-CoV-2 plays a crucial role in entering viruses into the host cell by binding to angiotensin-converting enzyme 2 (ACE-2), and this specific interaction represents a promising drug target for the identification of potential drugs. This study aimed at the receptor-binding domain of S protein (RBD of nCoV-SP) and the ACE-2 receptor as a promising target for developing drugs against SARS-CoV-2. Over 100 different flavonoids with antioxidant, anti-inflammatory, and antiviral properties from different literatures were taken as a ligand or inhibitor for molecular docking against target protein RBD of nCoV-SP and ACE-2 using PyRX and iGEMDOCK. Top flavonoids based on docking scores were selected for the pharmacokinetic study. Selected flavonoids (hesperidin, naringin, ECGC, and quercetin) showed excellent pharmacokinetics with proper absorption, solubility, permeability, distribution, metabolism, minimal toxicity, and excellent bioavailability. Molecular dynamics simulation studies up to 100 ns exhibited strong binding affinity of selected flavonoids to RBD of nCoV-SP and ACE-2, and the protein-ligand complexes were structurally stable. These identified lead flavonoids may act as potential compounds for developing effective drugs against SARS-CoV-2 by potentially inhibiting virus entry into the host cell.

"Identification of Nafamostat and VR23 as COVID-19 drug candidates by targeting 3CLpro and PLpro."

The sudden increase in the COVID-19 epidemic affected by novel coronavirus 2019 has jeopardized public health worldwide. Hence the necessities of a drug or therapeutic agent that heal SARS-CoV-2 infections are essential requirements. The viral genome encodes a large Polyprotein, further processed by the main protease/ 3C-like protease (3CLpro) and papain-like proteases (PLpro) into 16 nonstructural proteins to form a viral replication complex. These essential functions of 3CLpro and PLpro in virus duplication make these proteases a promising target for discovering potential therapeutic candidates and possible treatment for SARS-CoV-2 infection. This study aimed to screen a unique set of protease inhibitors library against 3CLpro and PLpro of the SARS-CoV-2. A molecular docking study was performed using PyRx to reveal the binding affinity of the selected ligands and molecular dynamic simulations were executed to assess the three-dimensional stability of protein-ligand complexes. The pharmacodynamics parameters of the inhibitors were predicted using admetSAR. The top two ligands (Nafamostat and VR23) based on docking scores were selected for further studies. Selected ligands showed excellent pharmacokinetic properties with proper absorption, bioavailability and minimal toxicity. Due to the emerging and efficiency of remdesivir and dexamethasone in healing COVID-19 patients, ADMET properties of the selected ligands were thus compared with it. MD Simulation studies up to 100 ns revealed the ligands' stability at the target proteins' binding site residues. Therefore, Nafamostat and VR23 may provide potential treatment options against SARS-CoV-2 infections by potentially inhibiting virus duplication though more research is warranted.

Evaluation of potential drugs against leishmaniasis targeting catalytic subunit of Leishmania donovani nuclear DNA primase using ligand based virtual screening, docking and molecular dynamics approaches.

Leishmania donovani, causes leishmaniasis, a global health trouble with around 89 different countries and its population under its risk. Replication initiation events have been instrumental in regulating the DNA duplication and as the small subunit of L. donovani nuclear DNA primase (Ld-PriS) inherits the catalytic site, it plays a vital role in DNA replication. In this study we have aimed Ld-PriS for the first time as a prospective target for the application of drug against Leishmania parasite. 3-D structures of Ld-PriS were built and ligand-based virtual screening was performed using hybrid similarity recognition techniques. Ligands from the ZINC database were used for the screening purposes based on known DNA primase inhibitor Sphingosine as a query. Top 150 ligands were taken into consideration for molecular docking against the query protein (Ld-PriS) using PyRx and iGEMDOCK softwares. Top five compounds with the best docking score were selected for pharmacokinetic investigation and molecular dynamic simulation. These top five screened inhibitors showed very poor binding affinity toward the catalytic subunit of human primase indicating their safety toward the host normal replication mechanism. The top five compounds showed good pharmacokinetic profiles and ADMET predictions revealed good absorption, solubility, permeability, uniform distribution, proper metabolism, minimal toxicity and good bioavailability. Simulation studies upto 50 ns revealed the three leads ZINC000009219046, ZINC000025998119 and ZINC000004677901 bind with Ld-PriS throughout the simulation and there were no huge variations in their backbone suggesting that these three may play as potential lead compounds for developing new drug against leishmaniasis.Communicated by Ramaswamy H. Sarma.

Identification of potential inhibitors against SARS-CoV-2 by targeting proteins responsible for envelope formation and virion assembly using docking based virtual screening, and pharmacokinetics approaches.

WHO has declared the outbreak of COVID-19 as a public health emergency of international concern. The ever-growing new cases have called for an urgent emergency for specific anti-COVID-19 drugs. Three structural proteins (Membrane, Envelope and Nucleocapsid protein) play an essential role in the assembly and formation of the infectious virion particles. Thus, the present study was designed to identify potential drug candidates from the unique collection of 548 anti-viral compounds (natural and synthetic anti-viral), which target SARS-CoV-2 structural proteins. High-end molecular docking analysis was performed to characterize the binding affinity of the selected drugs-the ligand, with the SARS-CoV-2 structural proteins, while high-level Simulation studies analyzed the stability of drug-protein interactions. The present study identified rutin, a bioflavonoid and the antibiotic, doxycycline, as the most potent inhibitor of SARS-CoV-2 envelope protein. Caffeic acid and ferulic acid were found to inhibit SARS-CoV-2 membrane protein while the anti-viral agent's simeprevir and grazoprevir showed a high binding affinity for nucleocapsid protein. All these compounds not only showed excellent pharmacokinetic properties, absorption, metabolism, minimal toxicity and bioavailability but were also remain stabilized at the active site of proteins during the MD simulation. Thus, the identified lead compounds may act as potential molecules for the development of effective drugs against SARS-CoV-2 by inhibiting the envelope formation, virion assembly and viral pathogenesis.

Linking gut microbiota with the human diseases.

The human gut is rich in microbes. Therefore, it is of interest to document data to link known human diseases with the gut microbiota. Various factors like hormones, metabolites and dietary habitats are responsible for shaping the microbiota of the gut. Imbalance in the gut microbiota is responsible for the pathogenesis of various disease types including rheumatoid arthritis, different types of cancer, diabetes mellitus, obesity, and cardiovascular disease. We report a review of known data for the correction of dysbiosis (imbalance in microbe population) towards improved human health.

Comparative evaluation of dexmedetomidine and esmolol on hemodynamic responses during laparoscopic cholecystectomy.

BACKGROUND: The advent of laparoscopic surgery has benefited the patient and surgeon; however creation of pneumoperitoneum for same has bearings during the perioperative period. These effects of pneumoperitoneum are associated with significant haemodynamic changes, increasing the morbidity of the patient. AIM: The present study compared the efficacy of dexmedetomidine and esmolol on hemodynamic responses during laparoscopic cholecystectomy Materials and Methods: A total of 90 patients aged 20-60 y, American Society of Anaesthesiologists (ASA) physical status I or II, of either sex, planned for laparoscopic cholecystectomy were included. The patients were randomly divided into three groups of 30 each. Group D received dexmedetomidine loading dose 1 mcg/kg over a period of 15 min and maintenance 0.5 mcg/kg/h throughout the pneumoperitoneum. Group E received esmolol loading dose 1 mg/kg over a period of 5 min and maintenance 0.5 mg/kg/h throughout the pneumoperitoneum. Group C received same volume of normal saline. MEASUREMENTS: Heart rate (HR), systolic blood pressure, diastolic blood pressure and mean arterial pressure (MAP) were recorded preoperative, after study drug, after induction, after intubation, after pneumoperitoneum at 15 min intervals, post pneumoperitoneum and postoperative period after 15 min. Propofol induction dose, intraoperative fentanyl requirement and sedation score were also recorded. RESULTS: In group D, there was no statistically significant increase in HR and blood pressure after pneumoperitoneum at any time intervals, whereas in Group E, there was a statistical significant increase in MAP after pneumoperitoneum at 15, 45, and 60 min only and HR during the whole pneumoperitoneum period. There was a significant decrease in induction dose of propofol and intraoperative fentanyl requirement in Group D and E, compared to Group C (p<0.0001). CONCLUSION: Dexmedetomidine is more effective than esmolol for attenuating the hemodynamic response to pneumoperitoneum in elective laparoscopic cholecystectomy. Dexmedetomidine and esmolol also reduced requirements of anaesthetic agents.

A highly basic sequence at the N-terminal region is essential for targeting the DNA replication protein ORC1 to the nucleus in Leishmania donovani.

The conserved eukaryotic DNA replication protein ORC1 is one of the constituents of pre-replication complexes that assemble at or very near origins prior to replication initiation. ORC1 has been shown to be constitutively nuclear in Leishmania major. This study investigates the sequences involved in nuclear localization of ORC1 in Leishmania donovani, the causative agent of visceral leishmaniasis. Nuclear localization signals (NLSs) have been reported in only a few Leishmania proteins. Functional analyses have delineated NLSs to regions of ~60 amino acids in length in the tyrosyl DNA phosphodiesterase I and type II DNA topoisomerase of L. donovani, and in the L. major kinesin KIN13-1. Using a panel of site-directed mutations we have identified a sequence essential for nuclear import of LdORC1. This sequence at the N terminus of the protein comprises residues 2-5 (KRSR), with K2, R3 and R5 being crucial. Independent mutation of the K2 residue causes exclusion of the protein from the nucleus, while mutating the R5 residue leads to diffusion of the protein throughout the cell. This sequence, however, is insufficient for targeting a heterologous protein (ß-galactosidase) to the nucleus. Analysis of additional ORC1 mutations and reporter constructs reveals that while the highly basic tetra-amino acid sequence at the N terminus is essential for nuclear localization, the ORC1 NLS in its entirety is more complex, and of a distributive character. Our results suggest that nuclear localization signalling sequences in Leishmania nuclear proteins are more complex than what is typically seen in higher eukaryotes.