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Tubulin protein is the fundamental unit of microtubules, and comprises of α and β subunits arranged in an alternate manner forming protofilaments. These longitudinal protofilaments are made up of intra- (α-β) and inter-dimer (β-α) interactions. Literature review confirms that GTP hydrolysis results in considerable structural rearrangement within GTP binding site of β-α dimer interface after the release of γ phosphate. In addition to this, the intra-dimer interface exhibits structural rigidity which needs further investigation. In this study, we explored the reasons for the flexibility and the rigidity of the β-α dimer and the α-β dimer respectively through molecular simulation and Anisotropic Normal Mode based analysis. As per the sequence alignment report, two glycine residues (Gly96 and Gly98) were observed in the T3 loop of the β subunit which get substituted by Asp98 and Ala100 in the T3 loop of the α subunit. The higher mobility of glycine residues contributes to the flexibility of the T3 loop of inter-dimer when they come in direct contact with the GTPase Activating Protein (GAP) domain of the subunit. This was confirmed through RMSD, RMSF and Radius of Gyration based studies. Conversely, the intra-dimer exhibited a lower mobility in the absence of glycine residues. As per ANM based analysis, positive domain correlations were observed between T3 loop and GAP domain of intra- and inter- dimeric contact regions. However, these correlation motions were higher in the intra-dimer as compared to the inter-dimer interface. Thus on the basis of our findings, we hypothesize that the higher flexibility of T3 loop and the GAP domain of the inter-dimer is required for structural rearrangement and protofilament stability during hydrolysis. Furthermore, the slightly rigid nature of the T3 loop and the GAP domain of the intra-dimer assists in enhancing the monomer-monomer interaction through the higher positive domain correlation.

Fanconi anemia (FA) is a rare genetic disorder characterized by bone marrow failure, predisposition to cancer, and congenital abnormalities. FA is caused by pathogenic variants in any of 22 genes involved in the DNA repair pathway responsible for removing interstrand crosslinks. FANCL, an E3 ubiquitin ligase, is an integral component of the pathway, but patients affected by disease-causing FANCL variants are rare, with only nine cases reported worldwide. We report here a FANCL founder variant, anticipated to be synonymous, c.1092G>A;p.K364=, but demonstrated to induce aberrant splicing, c.1021_1092del;p.W341_K364del, that accounts for the onset of FA in thirteen cases from South Asia, twelve from India and one from Pakistan. We comprehensively illustrate the pathogenic nature of the variant, provide evidence for a founder effect, and propose including this variant in genetic screening of suspected FA patients in India and Pakistan, as well as those with ancestry from these regions of South Asia.

The case study unveils the likely mechanism of a novel stop-loss DAX1 variant preceding the prolonged precocious puberty in the adrenal hypoplasia congenital (AHC) boy. A boy aged five years and nine months initially examined for the primary adrenal insufficiency symptoms. Next-generation sequencing confirmed the X-linked inheritance of a novel stop-loss DAX1 variant: c.1411T>C/p.Ter471Gln associated with AHC in the patient. The patient was subjected to a brief clinical follow-up from 11 to 15.1 years of age. The effect of the mutant-DAX1 variant (p.Ter471Gln) on DAX1-steroidogenic factor 1 (SF1) (protein-protein) interaction was studied by protein-protein docking using the ClusPro-online tool. At 5.9 yrs of age, the patient exhibited precocious puberty with the secondary sexual characteristics of Tanner 2 stage (of 9–14 yrs of age). The patient showed primary adrenal insufficiency with diminished cortisol concentrations at blood serum (25 ng/ml) and urine (3.55 μg/24 h) levels. Upon steroidal exposure, the patient showed normalized serum cortisol levels of 45–61 ng/ml. However, the precocious puberty got prolonged with the increased penis length of 8.5 cm and the bone age of 18 yrs old during the follow-up. The patient showed increased basal serum adrenocorticotropic hormone (110->2000 pg/ml) and follicle-stimulating hormone (18.4–22.3 mIU/ml) concentrations. Following an elevated hypothalamic-pituitary-gonadal axis activity witnessed upon gonarellin stimulation. Protein-protein docking confirmed a weaker interaction between the mutant-DAX1 (p.Ter471Gln) protein and the wild-SF1 protein. Overall, we hypothesize the weakened mutant-DAX1-SF1 (protein-protein) interaction could govern the prolonged precocious puberty augmented with the elevated hypothalamic-pituitary-gonadal/adrenal axis responses via SF1-induced neuronal nitric oxide synthetase activation in the patient.

Background: Human β tubulin displays resistance to drugs like Taxol and Vinblastine due to amino acids substitutions within and outside the drug binding site. Objective: This study focuses on the effect of amino acid substitutions outside the drug binding site on drug resistance. Amino acid substitution like R306C (mut2) is associated with Taxol resistance and D197N (mut1) and K350N (mut3) are associated with Vinblastine resistance. However, the mechanism of resistance has not been understood yet. This study has attempted to investigate the mechanism of resistance. Methods: SWISSMODEL server was used to model the wild and the mutant β subunits which were later considered for protein-protein and protein-ligand docking using HADDOCK and AutoDock 1.5.6 software respectively. Dimer mutants were generated using Swisspdbviewer. POCASA 1.1 server was used to calculate the overall effect of substitution on pocket volume and the effect of substitution on domain mobility was explored using GROMACS software. Results: From sequence perspective, amino acid replacement in all three positions viz. D197N (mut1), R306C (mut2) and K350N (mut3) were found to have a deleterious effect on the stability of the protein. This study was further confirmed through structural analysis. Change in hydrogen bonding pattern was observed within the site of substitution in modeled mut1 and mut3 which is known to be specifically involved in Vinblastine interaction. In mut2 associated with Taxol binding, the hydrogen bonding pattern remained unaltered. All three mutants showed better protein-protein (β-β) interactions compared to the wild-type. Pocket size analysis in β subunit revealed that Taxol binding site increased in size after substitution in mut2 compared to the wild-type. However, the size of the Vinblastine binding site in the dimer interface remained the same before and after the substitution in wild and the mutants. Wild-type (β monomer and αβ dimer) associated with Taxol and Vinblastine, respectively showed better drug interaction compared to their mutants. Conclusion: This study throws light on the mechanism of drug resistance due to amino acid substitutions outside the drug binding site. It was found that amino acid substitution outside the drug site enhanced protein-protein interaction between the β-β subunits.

In this work, the binding mode of antibiotic drug kanamycin with Calf thymus DNA (ctDNA) was assessed using thermodynamic, spectroscopic and molecular docking studies. Isothermal Titration Calorimetry (ITC) studies suggest that the binding is enthalpically favored with a small entropic change and a binding constant of the order of 104 mol−1 dm3, which remains almost same in the studied temperature range. Studies in the presence of osmolytes tetraethylene glycol and sucrose show no appreciable change in the binding behaviour as well as on the stability of the DNA-kanamycin complex. These finding suggests that the number of water molecule released or taken up is not significant in the binding process. Competitive fluorescence displacement assay and circular dichroism studies show that kanamycin follows groove binding mode of interaction. Molecular docking studies also supports that kanamycin prefers groove binding mode, stabilized by hydrogen bonding.

Background and aims: Potentially involved proteins which are implicated as a specific target for any diseased condition mayimplicate certain unusual features in several pathological conditions. Human Matrix metalloproteinase (MMP) family ofendopeptidases is one such family responsible for many beneficial as well as several pathological critical diseases. With the adventof field of bioinformatics and computational efforts can aid researchers to comprehend their system of work. Methodology: Aninsilico characterization of the MMP family has been carried out to analyze their primary, secondary, structural and functionalperspective . The research has been focused on specific MMPs in which the further study was based on Mutational analysisconfirming the pathogenicity of MMPs in cancer metastasis. The basic approach was to screen large protein families which playsdual role during normal and diseased conditions. Results: Thus it is hypothesized that cysteine rich and highly thermostableMMPs might be key players in diseased conditions. Conclusion: It can also be concluded that the disease responsive MMPs mightbe considered as promising targets for treatment of cancer.

Background The marketed synthetic (Dipeptidyl peptidase-IV) DPP-IV Inhibitors are expensive antidiabetic drugs and have been reported to cause unacceptable adverse effects such as pancreatitis, angioedema, thyroid and pancreatic cancers. In this scenario research to develop novel DPP-IV Inhibitors from alternative sources is the need of the hour. Hypothesis/purpose Terminalia arjuna, a medicinal herb with antidiabetic and cardioprotective activities may represent a natural DPP-IV Inhibitor, the DPP-IV Inhibitory activity of which may translate into demonstrable therapeutic benefits in setting of diabetes with cardiovascular co-morbidities. Study design The study type used for the present study was an experimental (In vitro, In vivo and In silico) design. Method The DPP-IV Inhibitory, antidiabetic and cardioprotective effects of Terminalia arjuna was evaluated in the experimental model of myocardial infarction co-existing with diabetes. To determine the active principle of Terminalia arjuna responsible for DPP-IV Inhibitory activity, the crystal structure of DPP-IV was considered as receptor which was docked against Arjunetin, Arjungenin, Arjunic acid, Arjunone, Ellagic acid, Gallic acid, Sitagliptin and Vildagliptin. The binding sites as well as affinity of various active ingredients of Terminalia arjuna for DPP- IV enzyme was elucidated using in silico studies and compared to Vildagliptin. Results Terminalia arjuna demonstrated significant DPP-IV Inhibitory, antidiabetic (significant reduction in HbA1C) and cardioprotective effects (restoration of myocardial CPK-MB) in the experimental model of myocardial infarction co-existing with diabetes. The cardioprotective efficacy correlated to its DPP-IV Inhibitory activity. The active ingredients of Terminalia arjuna (Arjunetin, Arjungenin, Arjunic Acid Arjunone, Ellagic acid and Gallic acid) demonstrated significant inhibition of DPP-IV enzyme. Arjunic acid and Arjunone prefers the active site pocket of DPP-IV enzyme. Compounds like Arjunetin and Vildagliptin prefers to bind near the interface region of the DPP-IV as their biological active forms are homodimer. Sitagliptin binds near the α/β hydrolase domain. Conclusion The DPP-IV Inhibitory activity of Terminalia arjuna was found to be comparable to Vildagliptin. The DPP-IV Inhibitory activity translated into significant cardioprotective effects in the setting of diabetes. The active ingredient of Terminalia arjuna; Arjunetin, Arjungenin, Ellagic acid and Arjunic acid showed superior DPP-IV Inhibitory activity as compared to synthetic DPP-IV inhibitors (Sitagliptin and Vildagliptin) based on results of docking studies.

The most common characteristics of diverse age-related neurodegenerative diseases are aggregation and accumulation of the misfolded protein in the brain. Alzheimer׳s disease (AD) is one of these protein conformational diseases. Extracellular accumulation of amyloid β (Aβ) is one the neuropathological hallmarks of Alzheimer disease. Various studies have shown that mutation in specific hydrophobic region of Aβ protein inhibit the formation of β sheet, thus aggregation of this protein is stalled. The identification of such mutation in Aβ protein can help us in elucidating the etiology of sporadic Aβ. In our study we have selected three positions: 19ILU, 21ALA and 41ILU in Aβ protein based on their hydrophobic nature and substituted them with PRO ( βSheet breaker). The effects of the substitutions were analysed using molecular dynamics simulation studies. The results validated that the mutations in the specified regions change the hydrophobicity of the protein and the βsheet formation was declined to zero per cent.

Tubulin has received more attention as potent anticancer drug target because it is the fundamental unit of microtubules and plays an active role in cell division. The drugs against �-tubulin are mainly derived from terrestrial plants and marine resources. Of these, red and brown algae from the marine environment produce better secondary metabolites. These compounds are found to be active against cancer cell lines. Since there is no study reported till date about the activity of these compounds against human � -tubulin, we have investigated the role of 517 compounds available in the seaweed secondary metabolites database against modeled human � -tubulin. All the conformers of lead compounds (RL381, RL366, RL376 and RG012) when docked at the taxol binding site showed better interactions with the H1-S2 loop and M-loop which are actively involved in lateral interactions of tubulins and also with the helix H7 which is the connecting link between N-terminal and intermediate domain. Important residues involved in polar interaction by these lead compounds were D224, H227, R276, R282 and R359 which closely mimicked the interaction of taxol with � -tubulin. We then attempted to calibrate the available molecules based on their Lipinski rule, binding affinity and other descriptor based comparison to identify potential lead molecules which could be used as drugs against humantubulin.

Human Immunodeficiency Virus (HIV-1) poses a serious threat to the developing world and sexual transmission continues to be the major source of new infections. Therefore, the development of molecules, which prevent new HIV-1 infections, is highly warranted. In the present study, a panel of human hemoglobin (Hb)-α subunit derived peptides and their analogues, with an ability to bind gp120, were designed in-silico and their anti-HIV-1 activity was evaluated. Of these peptides, HbAHP-25, an analogue of Hb-α derived peptide, demonstrated significant anti-HIV-1 activity. HbAHP-25 was found to be active against CCR5-tropic HIV-1 strains (ADA5 and BaL) and CXCR4-tropic HIV-1 strains (IIIB and NL4-3). Surface plasmon resonance (SPR) and ELISA revealed direct interaction between HbAHP-25 and HIV-1 envelope protein, gp120. The peptide prevented binding of CD4 to gp120 and blocked subsequent steps leading to entry and/or fusion or both. Anti-HIV activity of HbAHP-25 appeared to be specific as it failed to inhibit the entry of HIV-1 pseudotyped virus (HIV-1 VSV). Further, HbAHP-25 was found to be non-cytotoxic to TZM-bl cells, VK2/E6E7 cells, CEM-GFP cells and PBMCs, even at higher concentrations. Moreover, HbAHP-25 retained its anti-HIV activity in presence of seminal plasma and vaginal fluid. In brief, the study identified HbAHP-25, a novel anti-HIV peptide, which directly interacts with gp120 and thus has a potential to inhibit early stages of HIV-1 infection.