Understanding a point mutation signature D54K in the caspase activation recruitment domain of NOD1 capitulating concerted immunity via atomistic simulation.

Point mutation D54K in the human N-terminal caspase recruitment domain (CARD) of nucleotide-binding oligomerization domain -1 (NOD1) abrogates an imperative downstream interaction with receptor-interacting protein kinase (RIPK2) that entails combating bacterial infections and inflammatory dysfunction. Here, we addressed the molecular details concerning conformational changes and interaction patterns (monomeric-dimeric states) of D54K by signature-based molecular dynamics simulation. Initially, the sequence analysis prioritized D54K as a pathogenic mutation, among other variants, based on a sequence signature. Since the mutation is highly conserved, we derived the distant ortholog to predict the sequence and structural similarity between native and mutant. This analysis showed the utility of 33 communal core residues associated with structural-functional preservation and variations, concurrently served to infer the cryptic hotspots Cys39, Glu53, Asp54, Glu56, Ile57, Leu74, and Lys78 determining the inter helical fold forming homodimers for putative receptor interaction. Subsequently, the atomistic simulations with free energy (MM/PB(GB)SA) calculations predicted structural alteration that takes place in the N-terminal mutant CARD where coils changed to helices (45 α3- L4-α4-L6- α683) in contrast to native (45T2-L4-α4-L6-T483). Likewise, the C-terminal helices 93T1-α7105 connected to the loops distorted compared to native 93α6-L7105 may result in conformational misfolding that promotes functional regulation and activation. These structural perturbations of D54K possibly destabilize the flexible adaptation of critical homotypic NOD1CARD-CARDRIPK2 interactions (α4Asp42-Arg488α5 and α6Phe86-Lys471α4) is consistent with earlier experimental reports. Altogether, our findings unveil the conformational plasticity of mutation-dependent immunomodulatory response and may aid in functional validation exploring clinical investigation on CARD-regulated immunotherapies to prevent systemic infection and inflammation.Communicated by Ramaswamy H. Sarma.

A comprehensive integrated gene network construction to explore the essential role of Notch 1 in lung adenocarcinoma (LUAD).

The heterogeneous biological landscape of non-small cell lung cancer (NSCLC) is largely attributed to the activation of Notch signalling pathway. Among the Notch family transmembrane proteins, neurogenic locus notch homolog protein1 (NOTCH1) is a putative oncogene in NSCLC which activates the pathway as negative prognostic factor. This study aims to explore integrated network approach in lung adenocarcinoma (LUAD) especially linked to the notch pathway and its receptors. Our gene set enrichment analysis reveals the key Notch pathway genes are predominantly down regulated in LUAD. There were 675 genes with a total of 6517 functional interactions and 6 densely connected clusters of 38 miRNAs, 84 transcription factors with 156 edges identified through network construction. Here we report five key genes namely NOTCH1, CDH1, ERBB2, GAPDH and COL1A1 significantly enriched in Notch pathway which are further validated through the KM plot, box plots, stage plots and TIMER analysis. In addition, the NOTCH1 receptor is strongly linked to the immune checkpoint inhibitor CD274 (PD-L1) and can be considered as prognostic marker and tumour suppressor gene in LUAD which surely provide the basis for early diagnosis and futuristic immunotherapeutic targets for LUAD.Communicated by Ramaswamy H. Sarma.

Discovery of andrographolide hit analog as a potent cyclooxygenase-2 inhibitor through consensus MD-simulation, electrostatic potential energy simulation and ligand efficiency metrics.

Cyclooxygenase-2 (COX-2) is the key enzyme responsible for the conversion of arachidonic acid to prostaglandins that display pro-inflammatory properties and thus, it is a potential target protein to develop anti-inflammatory drugs. In this study, chemical and bio-informatics approaches have been employed to find a novel potent andrographolide (AGP) analog as a COX-2 inhibitor having better pharmacological properties than aspirin and rofecoxib (controls). The full amino acid sequenced human Alpha fold (AF) COX-2 protein (604AA) was selected and validated for its accuracy against the reported COX-2 protein structures (PDB ID: 5F19, 5KIR, 5F1A, 5IKQ and 1V0X) followed by multiple sequence alignment analysis to establish the sequence conservation. The systematic virtual screening of 237 AGP analogs against AF-COX-2 protein yielded 22 lead compounds based on the binding energy score (< - 8.0 kcal/mol). These were further screened out to 7 analogs by molecular docking analysis and investigated further for ADMET prediction, ligand efficiency metrics calculations, quantum mechanical analysis, MD simulation, electrostatic potential energy (EPE) docking simulation, and MM/GBSA. In-depth analysis revealed that AGP analog A3 (3-[2-[(1R,4aR,5R,6R,8aR)-6-hydroxy-5,6,8a-trimethyl-2-methylidene-3,4,4a,5,7,8-hexahydro-1H-naphthalen-1-yl]ethylidene]-4-hydroxyoxolan-2-one) forms the most stable complex with the AF-COX-2 showing the least RMSD value (0.37 ± 0.03 nm), a good number of hydrogen bonds (protein-ligand H-bond = 11, and protein H-bond = 525), minimum EPE score (- 53.81 kcal/mol), and lowest MM-GBSA before and after simulation (- 55.37 and - 56.25 kcal/mol, respectively) value compared to other analogs and controls. Thus, we suggest that the identified A3 AGP analog could be developed as a promising plant-based anti-inflammatory drug by inhibiting COX-2.

Andrographolide-based potential anti-inflammatory transcription inhibitors against nuclear factor NF-kappa-B p50 subunit (NF-κB p50): an integrated molecular and quantum mechanical approach.

The unregulated activation of nuclear factor-κB (NF-κB) is a critical event in the progression of various inflammatory diseases such as ulcerative colitis, asthma, rheumatoid arthritis, bacterial induced gastritis, etc. Hence, blocking the transcriptional activity of NF-κB is a promising strategy towards the development of an anti-inflammatory agent. In this study, an integrated molecular and quantum mechanical approach was carried out to find a new potent andrographolide (AGP)-based analog that can inhibit DNA binding to NF-κB p50 and manifest anti-inflammatory activity. Our approach includes multiple sequence alignment, virtual screening, molecular docking (protein-ligand and protein-DNA), in silico site-directed mutagenesis, ADMET prediction, DFT (HOMO, LUMO, HLG, and EPM energy) analysis, MD simulation, and MM/GBSA rescoring. The virtual screening analysis of 237 AGP analogs yielded the five lead compounds based on the binding affinity. Further, molecular interactive docking and ADMET prediction of hit analogs revealed that Ana2 ((3Z,4S)-3-[2-[(4aR,6aS,7R,10aS,10bR)-3,3,6a,10b-tetramethyl-8-methylidene-1,4a,5,6,7,9,10,10a-octahydronaphtho[2,1-d][1,3]dioxin-7-yl]ethylidene]-4-hydroxyoxolan-2-one) is the most potent moiety as it displays the strongest binding affinity and better molecular/pharmacokinetic features. Moreover, DFT, MD simulation, and MM/GBSA studies corroborated the docking results and demonstrated better chemical and dynamic stability with the least binding free energy (- 29.99 kcal/mol) for the Ana2. Site-directed mutagenesis investigation (Cys62Ala) establishes the importance of the Cys62 amino acid residue towards the binding interaction and stability of Ana2 with NF-κB p50. Overall, the identified NF-κB p50 inhibitor opens up a new research horizon towards the development of plant-based anti-inflammatory drugs to combat progressive inflammatory diseases. Supplementary Information: The online version contains supplementary material available at 10.1007/s13205-022-03431-9.

Transcriptomic analysis of castration, chemo-resistant and metastatic prostate cancer elucidates complex genetic crosstalk leading to disease progression.

Prostate adenocarcinoma, with its rising numbers and high fatality rate, is a daunting healthcare challenge to clinicians and researchers alike. The mainstay of our meta-analysis was to decipher differentially expressed genes (DEGs), their corresponding transcription factors (TFs), miRNAs (microRNA) and interacting pathways underlying the progression of prostate cancer (PCa). We have chosen multiple datasets from primary, castration-resistant, chemo-resistant and metastatic prostate cancer stages for investigation. From our tissue-specific and disease-specific co-expression networks, fifteen hub genes such as ACTB, ACTN1, CDH1, CDKN1A, DDX21, ELF3, FLNA, FLNC, IKZF1, ILK, KRT13, KRT18, KRT19, SVIL and TRIM29 were identified and validated by molecular complex detection analysis as well as survival analysis. In our attempt to highlight hub gene-associated mutations and drug interactions, FLNC was found to be most commonly mutated and CDKN1A gene was found to have highest druggability. Moreover, from DAVID and gene set enrichment analysis, the focal adhesion and oestrogen signalling pathways were found enriched which indicates the involvement of hub genes in tumour invasiveness and metastasis. Finally by Enrichr tool and miRNet, we identified transcriptional factors SNAI2, TP63, CEBPB and KLF11 and microRNAs, namely hsa-mir-1-3p, hsa-mir-145-5p, hsa-mir-124-3p and hsa-mir-218-5p significantly controlling the hub gene expressions. In a nutshell, our report will help to gain a deeper insight into complex molecular intricacies and thereby unveil the probable biomarkers and therapeutic targets involved with PCa progression.

Identification of metal binding motifs in protein frameworks to develop novel remediation strategies for Hg2+ and Cr(VI).

Amino acid sequences in metal-binding proteins with chelating properties offer exciting applications in biotechnology and medical research. To enhance their application in bioremediation studies, we explicitly aimed to identify specific metal-binding chelating motifs in protein structures for two significant pollutants, such as mercury (Hg2+) and chromium Cr(V1). For this purpose, we have performed an extensive coordination chemistry approach by retrieving Hg2+ and Cr(V1) binding protein structures from the protein database and validated using the B-factor, a term defining uncertainty of the atoms and with occupancy to obtain the best binding motifs. Our analysis revealed that acidic amino acids like aspartic acid, glutamic acid, and basic amino acids such as cysteine and histidine are predominant in coordinating with these metals. The order of preference in Hg2+-bound structures is predicted to be Cys > His > Asp > Glu, and for Cr(V1) is His > Asp > Glu. Examination of the atomic coordinates and their distance from each metal revealed that the sulfur atoms of cysteine showing more preference towards Hg2+coordination with an atomic distance ranging from 1.5 to 2.9 Å. Likewise, oxygen atoms of aspartic acid, glutamic acid and nitrogen atoms of histidine are within 2 Å of Cr(V1) coordination. Based on these observations, we obtained C-C-C, C-X(2)-C-C-(X)2-C, H-C-H motifs for Hg2+, and D-X(1)-D, H-X(3)-E motif for Cr(V1) to be shared within the coordination space of 3 Å. As a future scope, we propose that the identified metal-binding chelating motifs are oligopeptides and can display on the surface of microorganisms such as Escherichia coli and Saccharomyces cerevisiae for effective removal of natural Hg2+ and Cr(V1) through biosorption. Hence, our results will provide the basis for futuristic bioremediation.

Delineating the folding perturbations and molecular mechanisms of Thr-Ala 642 mutation in Rab-GTPase activating protein Akt substrate of 160kDa and its impact on the aetiology of diabetes.

Diabetes Mellitus is a complex metabolic disorder with one of the highest prevalence rates in the world. The present study probes into the Thr-Ala 642 mutation of Akt substrate of 160 kDa (AS160) which has been implicated in diabetes by the dysregulation of glucose transported vesicle 4 (GLUT4) translocation. Our study provides a possible evidence on structural basis dysfunction of AS160 and how the association of phosphorylated AS160 with 14-3-3, a downstream binding partner regulating GLUT4 translocation got disrupted due to T642A mutation. We initially derived the disease-causing mutation (Thr642Ala) among others through in-silico based statistical analysis. Subsequently, we interpreted the perturbation induced in the structural arrangement and their impaired interaction in core regions. Due to mutation, the key interfacial interactions between AS160-14-3-3 were changing from Thr642-Asp756, Thr642-Asp757, and Thr642-Lys659 for phosphorylated form, to Ala642-Val681 for mutant. Further, for phosphorylated AS160 the hotspot residues observed were Glu-629, Gln-635, His-641, Lys-653 and Arg-842 which changed to Arg-637, His-641 for mutant. Eventually, the molecular dynamics analysis revealed that local region for phosphorylation site of AS160 is reducing the flexibility, whereas mutation is making the region more flexible. Principal component analysis and Free energy landscape analysis together reveals phosphorylated AS160 is occupying less phase space with more stable landscape when compared to mutant. Our study strongly confers the destabilizing effect of the point mutation at a conserved site providing novel insights right down to the residual level of the conformational dysregulation of AS160 implicating deadly disease.

Exploring the structural significance of molecular switch mechanism alias motif phosphorylation in Wnt/ß-catenin and their crucial role in triple-negative breast cancer.

ß-Catenin, a key transcriptional factor involved in the canonical Wnt signaling pathway, is regulated by a cascade of phosphorylations and plays a major role in the progression of triple-negative breast cancer (TNBC). However, the phosphorylation induced conformational changes in a ß-Catenin is still poorly understood. Hence, we adopted a conventional molecular dynamics approach to study phosphorylations present in a sequence motif Ser 552 675 and Tyr670 of the ß-Catenin domain and analyzed in terms of structural transitions, bond formation, and folding-misfolding conformations. Our results unveil the ß-Catenin linear motif 549-555 (RRTSMGG) of armadillo repeats domain prefers order to disorder state. In contrast, helix C associated with 670-678 (YKKRLSVEL) motif prefers disorder to order upon phosphorylation of Ser 552 675 and Tyr670. In addition, the crucial secondary structural transition from α-helix to coil induced by phospho Ser552 and phospho Tyr670 of ß-Catenin ARM domain connecting helix C modifies conformational diversity and binding affinities of the complex interaction in functional regulation significantly. Moreover, the post phosphorylation disrupted the hydrogen bond interactions (Ser552-Arg549, Arg550-Asp546 and Ser675-Lys672) and abolished the residual alliance with hydrophobic interactions (Tyr670-Leu674) that easily interrupt in secondary structure packing as well as folding conformations connecting ARM and helix C (R10, 12 & R1C) compared to unphosphorylation. Our integrated computational analysis may help in shedding light on understanding the induced folding and unfolding pattern due to motif phosphorylations. Overall, our results provide an atomistic structural description of the way phosphorylation facilitates conformational and dynamic changes in ß-Catenin, a fundamental molecular switch mechanism in triple-negative breast cancer pathogenesis.

Bioactivity of melianone against Salmonella and in silico prediction of a membrane protein target.

Melianone, the protolimonoid (24, 25-epoxyflindissone), was isolated from the medicinal tree species, Swietenia mahagoni (L.) JACQ (Meliaceae). The compound isolated from petroleum ether leaf extracts (5.39%) was quantified using high-performance thin-layer chromatography (HPTLC) method. In antimicrobial assays melianone inhibited Salmonella ser. Typhi with an MIC of 0.053 µM. Induced Fit Docking (IFD) of the ligand, melianone, with proteins involved in anaerobic virulence of the pathogen, revealed that it binds with FocA (a transport protein of formate ions) at its "periplasmic opening" with a glide energy of - 51.8576 kcal mol-1. Melianone altered the overall conformation of the protein (protomer A) by 0.347 Å RMSD. It induced a notable protein topology (Ω loop region) shift in the channel from an intermediate-open to a closed-state conformation and was supported by molecular dynamic simulations performed. FocA, a protein that contributes to its survival under anaerobic conditions, was further evaluated experimentally, after exposure of Salmonella ser. Typhi to melianone, resulting in the altered homeostasis of formate.

Structural debilitation of mutation G322D associated with MSH2 and their role in triple negative breast cancer.

The missense mutation in the mismatch repair gene MSH2 underlies in several hereditary cancers. In this study, we have detailed the disruptive mutation of G322D that overtly pathogenic and clinically relevant to the triple negative breast cancer (TNBC) on the basis of structural aspect to untangle the unknown factors. We systematically evaluated the conformational changes that undergo upon mutation from the annotation of intra-residual contacts, secondary structural arrangements and fold recognition through molecular dynamics simulation. At first, we interpreted the total of 88 intra-molecular interaction which is required minimally to maintain the native structural architecture. Adequately, the molecular dynamics approach is well contributed toward structural modification that takes places in C-terminal linking 290-294(L20), 339-347(L23-T16- α9-T17) and 373-395(α12-T19-20-L26-T21-L27-T22) in contrast to native 290-294(ß8), 339-347(T20-α9-T21-α10), 373-395(α12-T19-20-L26-T21-L27-T22) provides a straightforward evidence that is underpinning destabilization and protein misfolding. Eventually, we have highlighted the structural debilitation of G322D in the core region of 303-309 L23-T18-α6 - L21- α8, and 326-330 α7-T19-L25 -α9 notably the connecting elements of secondary structural propensity (loop-helix) in folding pack were completely abrupted which helps to keep native form. Essentially, the information gained in our study on residual interaction and conformational transitions in the structure of mutant MSH2 provides valuable insights to understand the clues of functional behavior and also pave the way to frame suitable and improved therapeutical targets.Communicated by Ramaswamy H. Sarma.

Streptozocin; a GLUT2 binding drug, interacts with human serum albumin at loci h6DOM3-h7DOM3.

Streptozocin (STZ) is a broad range antibiotic, highly genotoxic, antineoplastic and hyperglycemic. HSA is the most abundant protein in physiology and it binds to almost all exogenic and endogenic ligands, including drugs. STZ-induced fluorescence quenching of HSA has been done at pH 7.4, pH 3.5 and at pH 7.4 with 4.5 M urea at temperatures 286 K, 291 K, and 306 K. Ksv found to be 103 M-1, binding constant 1.5X103M-1 and binding sites ~1. But, Ksv for HSA and glucopyranose interaction was found lesser than that of HSA-STZ binding. Binding of STZ/glucopyranose on HSA seems to result in complex formation as calculated Kq > 1010 M-1 s-1. The number of binding sites, binding constants, and binding energies were increased with temperature. The ΔG0, ΔH0, and ΔS0 for HSA-STZ interaction were found to be -17.7 × 103 J·mol-1; 2.34 × 105 J·mol-1 and 841 JK-1 mol-1 respectively at pH 7.4 and 291 K. The comparative bindings of N, F and I states of HSA with STZ and their molecular docking analyses indicate that IIIA-B junction (i.e., inter-helix h6DOM3-h7DOM3) is the probable binding site, a locus close to fatty acid binding site-5. These results could be useful for therapeutic and analytical exploitation of STZ, as albumin used as the vehicle for drug delivery.

R516Q mutation in Melanoma differentiation-associated protein 5 (MDA5) and its pathogenic role towards rare Singleton-Merten syndrome; a signature associated molecular dynamics study.

Singleton-Merten syndrome, a critical and rare multifactorial disorder that is closely linked to R516Q mutation in MDA5 protein associated with an enhanced interferon response in the affected individual. In the present study, we provide conclusive key evidence on R516Q mutation and their connectivity towards sequence-structural basis dysfunction of MDA5 protein. Among the various mutations, we found R516Q is the most pathogenic mutation based on mutational signature Q-A-[RE]-G-R-[GA]-R-A-[ED]-[DE]-S-[ST]-Y-[TSAV]-L-V designed from our work. Further, we derived a distant ortholog for this mutational signature from which we identified 343 intra-residue interactions that fall communally in the position required to maintain the structural and functional integration of protein architecture. This identification served us to understand the critical role of hot spots in residual aggregation that holds a native form of folding conformation in the functional region. In addition, the long-range molecular dynamics simulation demarcated the residual dependencies of conformational transition in distinct regions (L29360-370α18, α19380-410L31, α21430-480L33-α22-L35 and α24510-520L38) occurring upon R516Q mutation. Together, our results emphasise that the dislocation of functional hot spots Pro229, Arg414, Val498, Met510, Ala513, Gly515 and Arg516 in MDA5 protein which is important for interior structural packing and fold arrangements. In a nutshell, our findings are perfectly conceded with other experimental reports and will have potential implications in immune therapeutical advancement for rare singleton-merten syndrome.

A molecular simulation analysis of vitamin D targets interleukin 13 (IL13) as an alternative to mometasone in asthma.

Asthma, a chronic lung disease characterized by obstruction of airway passage is characterized by inflammation and hyperresponsiveness with increase in the number of eosinophils. Interleukin-13, plays a significant role in causing inflammation during an asthmatic attack by bronchial constriction. Mometasone, a glucocorticoid has been used as the first line of administration for people affected with asthma for almost a decade. However, in several cases, people treated with mometasone have faced systemic and local side effects. To reduce these side effects, we hypothesized vitamin D that can be used as a substitute to mometasone. For this purpose, we employed the use of molecular docking and simulation studies for comparative study. The docking studies revealed the binding residues of interleukin-13 which are bound to the active site. Among all, we noticed three binding residue Leu83, His84 and Arg86 common for both mometasone and vitamin D. Also, the binding energies share a significant similarity between them. The docked complexes of mometasone and vitamin D with interleukin-13 were evaluated with molecular dynamics simulation. Consistently, the MD analysis uncovered the interesting note on conformational adaptation between the complexes as well as that vitamin D has the complementary binding efficiency to interleukin-13 as compared to mometasone. The substitution of vitamin D might provide a promising gateway to reduce the side effects caused by mometasone and also reduce the cost for treatment of asthma patients.

Integrative network-based approach identifies central genetic and transcriptomic elements in triple-negative breast cancer.

Triple-negative breast cancer (TNBC) has gained considerable attention as it oversteps about 15% of the deaths caused by breast cancer in women and is well known for its aggressive impact, high proliferation rate, intensity of tumor, and metastasis. In this study, we have given a new insight into TNBC, by introducing an integrative network methodology to understand the complexity of the candidate genes in TNBC. In the course of this analysis, the central network was constructed by refining the candidate genes present in the various databases. Eventually, the hub genes SRC, EGFR, JUN, CTNNB1, and MYC were derived using distinct topological parameters such as degree, betweenness centrality, closeness centrality, and clustering coefficient, which implicated a central role in TNBC. Furthermore, the identified hub genes were validated by utilizing molecular complex detection cluster analysis. A regulatory network was constructed to locate the transcriptional factors regulating these hub genes via MatInspector. Interestingly, we characterized that ZF02, MZF1, and PLAG are the common transcription factors which activate the hub genes. Moreover, a functional enrichment pathway was obtained from ClueGo, which depicted that all the hub genes have a significant role in toxicity-associated pathways as well as physiochemical featured pathways. In a nutshell, these prioritized genes could be the way out to uncover the molecular and therapeutic targets, leading to the development of personalized medicine for TNBC.

Evaluation of peptide designing strategy against subunit reassociation in mucin 1: A steered molecular dynamics approach.

Subunit reassociation in mucin 1, a breast cancer tumor marker, is reported as one of the critical factors for its cytoplasmic activation. Inhibition of its heterodimeric association would therefore result in loss of its function and alter disease progression. The present study aimed at evaluating peptide inhibitor designing strategies that may serve as antagonist against this receptor-ligand alliance. Several peptides and their derivatives were designed based on native residues, subunit interface, hydrogen bonding and secondary structure. Docking studies with the peptides were carried on the receptor subunit and their binding affinities were evaluated using steered molecular dynamics simulation and umbrella sampling. Our results showed that among all the different classes of peptides evaluated, the receptor based peptide showed the highest binding affinity. This result was concurrent with the experimental observation that the receptor-ligand alliance in mucin 1 is highly specific. Our results also show that peptide ligand against this subunit association is only stabilized through native residue inter-protein interaction irrespective of the peptide structure, peptide length and number of hydrogen bonds. Consistency in binding affinity, pull force and free energy barrier was observed with only the receptor derived peptides which resulted in favorable interprotein interactions at the interface. Several observations were made and discussed which will eventually lead to designing efficient peptide inhibitors against mucin 1 heterodimeric subunit reassociation.

Systematic prioritization of functional hotspot in RIG-1 domains using pattern based conventional molecular dynamic simulation.

BACKGROUND: Retinoic acid inducible gene 1 (RIG-1), multi-domain protein has a role-play in detecting viral nucleic acids and stimulates the antiviral response. Dysfunction of this protein due to mutations makes the route vulnerable to viral diseases. AIM: Identification of functional hotspots that maintains conformational stability in RIG-1 domains. METHODS: In this study, we employed a systematic in silico strategy on RIG-1 protein to understand the mechanism of structural changes upon mutation. We computationally investigated the protein sequence signature for all the three domains of RIG-1 protein that encloses the mutation within the motif. Further, we carried out a structural comparison between RIG-1 domains with their respective distant orthologs which revealed the minimal number of interactions required to maintain its structural fold. This intra-protein network paved the way to infer hotspot residues crucial for the maintenance of the structural architecture and folding pattern. KEY FINDINGS: Our analysis revealed about 40 hotspot residues that determine the folding pattern of the RIG-1 domains. Also, conventional molecular dynamic simulation coupled with essential dynamics provides conformational transitions of hot spot residues among native and mutant structures. Structural variations owing to hotspot residues in mutants again confirm the significance of these residues in structural characterization of RIG-1 domains. We believe our results will help the researchers to better comprehend towards regulatory regions and target-binding sites for therapeutic design within the pattern recognition receptor proteins. SIGNIFICANCE: Our protocol employed in this work describes a novel approach in identifying signature residues that would provide structural insights in protein folding.

Casting the critical regions in nucleotide binding oligomerization domain 2 protein: a signature mediated structural dynamics approach.

Nucleotide binding oligomerization domain 2 (NOD2), a protein involved in the first line defence mechanism has a pivotal role in innate immunity. Impaired function of this protein is implicated in disorders such as Blau syndrome and Crohn's disease. Since an altered function is linked to protein's structure, we framed a systematic strategy to interpret the structure-function relationship of the protein. Initiated with mutation-based pattern prediction and identified a distant ortholog (DO) of NOD2 from which the intra-residue interaction network was elucidated. The network was used to identify hotspots that serve as critical points to maintain the stable architecture of the protein. Structural comparison of NOD2 domains with a DO revealed the minimal number of intra-protein interactions required by the protein to maintain the structural fold. In addition, the conventional molecular dynamics simulation emphasized the conformational transitions at hot spot residues between native NOD2 domains and its respective mutants (G116R, R42W and R54A) structures. The analysis of intra-protein interactions globally and the displacement of residues locally around the mutational site revealed loss of several critical bonds and residues vital for the protein's function. Conclusively we report, about 10 residues in leucine-rich repeat, 13 residues in NOD and 6 residues in CARD domain are required by the NOD2 to maintain its function. This protocol will help the researchers to achieve for more prospective studies to attest druggable site utility in discovering novel drug candidates.

Screening of Furanone in Cucurbita melo and Evaluation of its Bioactive Potential Using In Silico Studies.

The work presented here attempts to screen for the presence of 4-Hydroxy-2,5-dimethyl-3(2H)-furanone in fruits and also any bioactive potential present in the fruit extracts. Curcurbita melo was selected for the study, and the fruit was crushed, filtered and extracted with ethyl acetate as the solvent. The resulting extract was subjected to disk diffusion test using Kirby Bauer method for checking its antimicrobial potential. Melon extract showed promising results against different clinical pathogens, 19-mm zone being the largest against Klebsiella pneumoniae and 17 mm against Shigella dysenteriae. GC-MS data of the melon extract confirmed the existence of furanone derivative in the extract. To evaluate the antimicrobial activity, in silico studies were performed using AutoDock 4.0 software, and topoisomerase was used as the target protein molecule for the isolated compound and 3-methyl-2-(2-oxopropyl)furan as the ligand. Further, the results were interpreted using PyMol and Ligplot plus softwares. The results confirmed the presence of molecular interactions between the protein molecule and the ligand. It can be envisaged that these interactions are responsible for the inhibitory effects of the extract .

Targeting inflammatory mediators with ferulic acid, a dietary polyphenol, for the suppression of monosodium urate crystal-induced inflammation in rats.

AIMS: The aim of this study was to investigate the anti-inflammatory effect of ferulic acid, a dietary phenol, on monosodium urate (MSU) crystal-induced inflammation in rats, an experimental model for acute gouty arthritis. For the purpose of comparison, colchicine was used as a reference drug. MAIN METHODS: Paw edema, levels/activities of elastase, lysosomal enzymes (acid phosphatase and ß-galactosidase), nitric oxide, lipid peroxidation, antioxidant status and pro-inflammatory cytokines (tumor necrosis factor alpha (TNF-α) and interleukin (IL)-1ß), and histology of ankle joints were evaluated in rats with MSU crystal-induced inflammation. The messenger RNA (mRNA) expression of pro-inflammatory cytokines (TNF-α and IL-1ß), NLRP3 (nucleotide oligomerization domain (NOD)-like receptor family, pyrin domain containing 3) inflammasomes, caspase-1, and the transcription factor nuclear factor kappa B p65 (NF-κB p65) was determined by real-time polymerase chain reaction (PCR) analysis. The protein expression of NF-κB p65 and TNF-α was detected by immunohistochemical analysis. Further, a molecular docking analysis was conducted to determine the ligand efficiency of ferulic acid towards NF-κB, apoptosis-associated speck-like protein containing a CARD (PYCARD/ASC), NLRP3, and pro-caspase-1. KEY FINDINGS: In the joint homogenate of rats with MSU crystal-induced inflammation, treatment with ferulic acid (30mg/kg body weight (b.wt)) decreased paw edema; the level/activity of elastase, lysosomal enzymes, nitric oxide, lipid peroxidation, and pro-inflammatory cytokines (TNF-α and IL-1ß); and the mRNA expression of NLRP3 inflammasomes, caspase-1, pro-inflammatory cytokines, and NF-κB p65. In addition, the protein expression of NF-κB p65 and TNF-α was also found to be significantly decreased. However, the antioxidant status (superoxide dismutase (SOD) and catalase (CAT)) were found to be increased. The molecular docking analysis showed that ferulic acid exhibited significant ligand efficiency towards pro-caspase-1, NF-κB, PYCARD/ASC, and NLRP3. SIGNIFICANCE: Our findings demonstrate the potential anti-inflammatory effect of ferulic acid on MSU crystal-induced inflammation in rats.

Ab initio coordination chemistry for nickel chelation motifs.

Chelation therapy is one of the most appreciated methods in the treatment of metal induced disease predisposition. Coordination chemistry provides a way to understand metal association in biological structures. In this work we have implemented coordination chemistry to study nickel coordination due to its high impact in industrial usage and thereby health consequences. This paper reports the analysis of nickel coordination from a large dataset of nickel bound structures and sequences. Coordination patterns predicted from the structures are reported in terms of donors, chelate length, coordination number, chelate geometry, structural fold and architecture. The analysis revealed histidine as the most favored residue in nickel coordination. The most common chelates identified were histidine based namely HHH, HDH, HEH and HH spaced at specific intervals. Though a maximum coordination number of 8 was observed, the presence of a single protein donor was noted to be mandatory in nickel coordination. The coordination pattern did not reveal any specific fold, nevertheless we report preferable residue spacing for specific structural architecture. In contrast, the analysis of nickel binding proteins from bacterial and archeal species revealed no common coordination patterns. Nickel binding sequence motifs were noted to be organism specific and protein class specific. As a result we identified about 13 signatures derived from 13 classes of nickel binding proteins. The specifications on nickel coordination presented in this paper will prove beneficial for developing better chelation strategies.