02932nas a2200313 4500008004100000022001400041245013200055210006900187260000800256520189100264653001602155653001302171653001102184653000902195653003302204653002302237653000902260653002102269100002202290700002502312700002302337700001902360700002602379700002702405700001702432700002002449700002502469856012402494 2020 eng d a0739-110200aDiscovery of potential multi-target-directed ligands by targeting host-specific SARS-CoV-2 structurally conserved main protease0 aDiscovery of potential multitargetdirected ligands by targeting  cMAY3 a

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection has resulted in the current COVID-19 pandemic. Worldwide this disease has infected over 2.5 million individuals with a mortality rate ranging from 5 to 10%. There are several efforts going on in the drug discovery to control the SARS-CoV-2 viral infection. The main protease (M-Pro) plays a critical role in viral replication and maturation, thus can serve as the primary drug target. To understand the structural evolution of M-Pro, we have performed phylogenetic and Sequence Similarity Network analysis, that depicted divergence of Coronaviridae M-Pro in five clusters specific to viral hosts. This clustering was corroborated with the comparison of M-Pro structures. Furthermore, it has been observed that backbone and binding site conformations are conserved despite variation in some of the residues. These attributes can be exploited to repurpose available viral protease inhibitors against SARS-CoV-2 M-Pro. In agreement with this, we performed screening of similar to 7100 molecules including active ingredients present in the Ayurvedic anti-tussive medicines, anti-viral phytochemicals and synthetic anti-virals against SARS-CoV-2 M-Pro as the primary target. We identified several natural molecules like delta-viniferin, myricitrin, taiwanhomoflavone A, lactucopicrin 15-oxalate, nympholide A, afzelin, biorobin, hesperidin and phyllaemblicin B that strongly binds to SARS-CoV-2 M-Pro. Intrestingly, these molecules also showed strong binding with other potential targets of SARS-CoV-2 infection like viral receptor human angiotensin-converting enzyme 2 (hACE-2) and RNA dependent RNA polymerase (RdRp). We anticipate that our approach for identification of multi-target-directed ligand will provide new avenues for drug discovery against SARS-CoV-2 infection. Communicated by Ramaswamy H. Sarma

10aCoronavirus10aCOVID-1910ahACE-210aMPro10amulti-target-directed ligand10aprotease inhibitor10aRdRp10aSARS-CoV-2 virus1 aJoshi, Rakesh, S.1 aJagdale, Shounak, S.1 aBansode, Sneha, B.1 aShankar, Shiva1 aTellis, Meenakshi, B.1 aPandya, Vaibhav, Kumar1 aChugh, Anita1 aGiri, Ashok, P.1 aKulkarni, Mahesh, J. uhttps://library.ncl.res.in/content/discovery-potential-multi-target-directed-ligands-targeting-host-specific-sars-cov-202420nas a2200265 4500008004100000245011300041210006900154260000800223300000600231490000700237520156200244653001401806653003101820653001401851653001301865653001301878653001101891653002401902100001901926700001901945700002601964700002201990700001902012856012302031 2021 eng d00aRepurposing the McoTI-II rigid molecular scaffold in to inhibitor of `papain superfamily' cysteine proteases0 aRepurposing the McoTIII rigid molecular scaffold in to inhibitor cJAN a70 v143 a

Clan C1A or `papain superfamily' cysteine proteases are key players in many important physiological processes and diseases in most living systems. Novel approaches towards the development of their inhibitors can open new avenues in translational medicine. Here, we report a novel design of a re-engineered chimera inhibitor Mco-cysteine protease inhibitor (CPI) to inhibit the activity of C1A cysteine proteases. This was accomplished by grafting the cystatin first hairpin loop conserved motif (QVVAG) onto loop 1 of the ultrastable cyclic peptide scaffold McoTI-II. The recombinantly expressed Mco-CPI protein was able to bind with micromolar affinity to papain and showed remarkable thermostability owing to the formation of multi-disulphide bonds. Using an in silico approach based on homology modelling, protein-protein docking, the calculation of the free-energy of binding, the mechanism of inhibition of Mco-CPI against representative C1A cysteine proteases (papain and cathepsin L) was validated. Furthermore, molecular dynamics simulation of the Mco-CPI-papain complex validated the interaction as stable. To conclude, in this McoTI-II analogue, the specificity had been successfully redirected towards C1A cysteine proteases while retaining the moderate affinity. The outcomes of this study pave the way for further modifications of the Mco-CPI design for realizing its full potential in therapeutics. This study also demonstrates the relevance of ultrastable peptide-based scaffolds for the development of novel inhibitors via grafting.

10aCathepsin10aClan CA cysteine proteases10acyclotide10acystatin10aMcoTI-II10apapain10aprotein engineering1 aMishra, Manasi1 aSingh, Vigyasa1 aTellis, Meenakshi, B.1 aJoshi, Rakesh, S.1 aSingh, Shailja uhttps://library.ncl.res.in/content/repurposing-mcoti-ii-rigid-molecular-scaffold-inhibitor-papain-superfamily-cysteine02369nas a2200289 4500008004100000022001400041245013700055210006900192260000800261300001400269490000700283520140400290653001901694653002501713653001501738653000601753653001801759653003101777653001901808100001901827700001901846700002601865700002201891700002401913700001901937856012301956 2022 eng d a0739-110200aCyclic peptide engineered from phytocystatin inhibitory hairpin loop as an effective modulator of falcipains and potent antimalarial0 aCyclic peptide engineered from phytocystatin inhibitory hairpin  cMAY a3642-36540 v403 a

Cystatins are classical competitive inhibitors of C1 family cysteine proteases (papain family). Phytocystatin superfamily shares high sequence homology and typical tertiary structure with conserved glutamine-valine-glycine (Q-X-V-X-G) loop blocking the active site of C1 proteases. Here, we develop a cysteine-bounded cyclic peptide (CYS-cIHL) and linear peptide (CYS-IHL), using the conserved inhibitory hairpin loop amino acid sequence. Using an in silico approach based on modeling, protein-peptide docking, molecular dynamics simulations and calculation of free energy of binding, we designed and validated inhibitory peptides against falcipain-2 (FP-2) and -3 (FP-3), cysteine proteases from the malarial parasite Plasmodium falciparum. Falcipains are critical hemoglobinases of P. falciparum that are validated targets for the development of antimalarial therapies. CYS-cIHL was able to bind with micromolar affinity to FP-2 and modulate its binding with its substrate, hemoglobin in in vitro and in vivo assays. CYS-cIHL could effectively block parasite growth and displayed antimalarial activity in culture assays with no cytotoxicity towards human cells. These results indicated that cyclization can substantially increase the peptide affinity to the target. Furthermore, this can be applied as an effective strategy for engineering peptide inhibitory potency against proteases.

10aCyclic peptide10afalcipain inhibitors10afalciparum10aP10aPhytocystatin10aprotein interaction mimics10aprotein&\#82111 aMishra, Manasi1 aSingh, Vigyasa1 aTellis, Meenakshi, B.1 aJoshi, Rakesh, S.1 aPandey, Kailash, C.1 aSingh, Shailja uhttps://library.ncl.res.in/content/cyclic-peptide-engineered-phytocystatin-inhibitory-hairpin-loop-effective-modulator02246nas a2200265 4500008004100000022001400041245007900055210006900134260000800203300001200211490000700223520141000230653002101640653000701661653002701668653001601695653002201711100002301733700002601756700002501782700002201807700002001829700002401849856010701873 2022 eng d a0022-284400aFunctional diversity of the lepidopteran ATP-binding cassette transporters0 aFunctional diversity of the lepidopteran ATPbinding cassette tra cAUG a258-2700 v903 a

The ATP-binding cassette (ABC) transporter gene family is ubiquitous in the living world. ABC proteins bind and hydrolyze ATP to transport a myriad of molecules across various lipid-containing membrane systems. They have been studied well in plants for transport of a variety of compounds and particularly, in vertebrates due to their direct involvement in resistance mechanisms against several toxic molecules/metabolites. ABC transporters in insects are found within large multigene families involved in the efflux of chemical insecticides and toxic/undesired metabolites originating from food and endogenous metabolism. This review deals with ABC transporter subfamilies of few agronomically important Lepidopteran pests. The transcriptional dynamics and regulation of ABC transporters during insect development emphasizes their functional diversity against insecticides, Cry toxins, and plant specialized metabolites. To generate insights about molecular function and physiological roles of ABCs, functional and structural characterization is necessary. Also, expansion and divergence of ABC transporter gene subfamilies in Lepidopteran insects needs more systematic investigation. We anticipate that newer methods of insect control in agriculture can benefit from an understanding of ABC transporter interactions with a vast range of natural specialized molecules and synthetic compounds.

10aABC transporters10aBt10ainsecticide resistance10aLepidoptera10aPlant metabolites1 aBarve, Pranoti, R.1 aTellis, Meenakshi, B.1 aBarvkar, Vitthal, T.1 aJoshi, Rakesh, S.1 aGiri, Ashok, P.1 aKotkar, Hemlata, M. uhttps://library.ncl.res.in/content/functional-diversity-lepidopteran-atp-binding-cassette-transporters02629nas a2200253 4500008004100000022001400041245013500055210006900190260000800259490000700267520172200274653001401996653002202010653002202032653003302054653002202087100002602109700001902135700001802154700002602172700003002198700001802228856012902246 2022 eng d a0739-110200aMolecular phylogeny, structure modeling and in silico screening of putative inhibitors of aerolysin of Aeromonas hydrophila EUS1120 aMolecular phylogeny structure modeling and in silico screening o cJUN0 v403 a

Aeromonas hydrophila, a Gram-negative bacterium, causes diseases in fish, resulting in excessive loss to the aquaculture industry. Aeromonas is a highly heterogeneous group of bacteria, and the heterogeneity of the genus is attributed to variation and diversity in the virulence factors and toxins among various Aeromonas strains. One of the major toxins aerolysin, secreted by the bacterium, causes hemorrhagic-septicemia and diarrhea and can serve as a drug target. Here, we describe characterization, molecular phylogeny, and homology modeling of the aerolysin of A. hydrophila strain EUS112 (Ah(EUS112)) cloned in our lab. The encoded aerolysin is 485 amino acids long with an N-terminal signal sequence of 23 amino acids. Phylogenetic analysis of the aerolysin of Ah(EUS112) revealed that it belongs to a diverse group of toxins, showing maximum similarity with aerolysins of other Aeromonas strains followed by Vibrio toxin. The homology model of the mature aerolysin of Ah(EUS112) was generated using the crystal structure of a mutant aerolysin (PDB\#3g4n) as the template, which showed that the encoded aerolysin exists as a channel protein. Validation of the generated model using bioinformatics tool confirmed it to be a good quality model that can be used for drug design. Molecular dock analysis revealed that drugs, aralia-saponin I, cyclamin, ardisiacrispin B, and aralia-saponin II bind to aerolysin with a higher affinity as compared to other drugs and at functionally important amino acids of aerolysin. Hence, these molecules can act as an effective therapeutics for inhibiting the aerolysin pore formation and curtail the severity of Aeromonas infection. Communicated by Ramaswamy H. Sarma

10aAerolysin10ahomology modeling10aMolecular docking10aMolecular dynamic simulation10aphylogenetic tree1 aYadav, Sunita, Kumari1 aPanwar, Deepak1 aSingh, Ankita1 aTellis, Meenakshi, B.1 aJoshi, Rakesh, Shamsunder1 aDixit, Aparna uhttps://library.ncl.res.in/content/molecular-phylogeny-structure-modeling-and-silico-screening-putative-inhibitors-aerolysin02364nas a2200229 4500008004100000022001400041245008700055210006900142260000900211300001200220490000700232520163500239653001201874653002301886653002501909653001101934653001401945100002601959700002401985700002202009856010302031 2023 eng d a0959-665800aRegulation of trehalose metabolism in insects: from genes to the metabolite window0 aRegulation of trehalose metabolism in insects from genes to the  cMAY  a262-2730 v333 a

Trehalose is a major circulatory sugar in the haemolymph of insects. It provides instant energy and protection against stress. Trehalose metabolism is associated with insect growth and development. The architecture and spatio-temporal expression dynamics of trehalose metabolism and transport genes are key for regulation. These genes are controlled by various transcription factors, largely linked to nutrition, insect development, and metamorphosis. Also, trehalose levels are affected by substrate affinities and modifications of enzymes involved in the pathway. A feedback mechanism involving the precursors and products can regulate trehalose metabolism. Further, the neuroendocrine system controls trehalose levels under normal and stressed conditions by producing different hormones. Hypotrehalosemic hormones work under surplus energy conditions to activate haemolymph trehalose uptake and degradation. In contrast, hypertrehalosemic hormones stimulate trehalose production in the fat body and its transport to the haemolymph. However, trehalose metabolism regulation in insects needs to be studied in detail. This review discusses aspects of trehalose synthesis, transport, and degradation dynamics in developmental transition and stress response. Unraveling the epigenetic factors, transcriptional control and chemical or genetic modulators can provide further insights into the intricate regulation of trehalose in a development- and tissue-specific manner. This molecular information about effectors and regulators of trehalose metabolism can be applied in developing diverse biotechnological applications.

10aHormone10aInsect development10aMetabolic regulation10aStress10atrehalose1 aTellis, Meenakshi, B.1 aKotkar, Hemlata, M.1 aJoshi, Rakesh, S. uhttps://library.ncl.res.in/content/regulation-trehalose-metabolism-insects-genes-metabolite-window02593nas a2200277 4500008004100000022001400041245012100055210006900176260000800245300001100253490000800264520165000272653002001922653002501942653001101967653002101978653001401999100002602013700003002039700002602069700002202095700002502117700002402142700002202166856012702188 2023 eng d a0378-111900aTrehalose transporter-like gene diversity and dynamics enhances stress response and recovery in Helicoverpa armigera0 aTrehalose transporterlike gene diversity and dynamics enhances s cAPR a1472590 v8623 a

Trehalose is a primary sugar and its distribution across the insect body, regulated by trehalose transporters (TRETs), is essential for sugar metabolism and energy homeostasis. The large diversity of Tret-like sugar trans-porters (ST), belonging to SLC2A transporter family, in polyphagous insects probably contributes to their extremely adaptive nature. We aim to study spatio-temporal expression dynamics and functional relevance of ST transcript variants in the lepidopteran model organism, Helicoverpa armigera. Identification of 69 putative Tret-like HaST transcript variants from databases and their digital gene expression analysis indicated tissue and development-specific expression patterns. Phylogenetic and sequence similarity network analysis of HaSTs signify evolutionary divergence, while motif and structure analysis depicted conserved signatures. In vitro gene expression validation for selected genes depicts that HaST09 and 69 are fat body and haemolymph-specific. While, HaST06, 30, 36 and 57 are developmental stage or sex-specific. HaST69 has high expression in the haemolymph of fifth instar larvae. In the presence of trehalose metabolism inhibitors and abiotic stress, HaSTs expression show dysregulation, indicating their possible association with trehalose metabolism and stress re-covery. In vivo gene silencing of HaST69 resulted in reduced trehalose accumulation in the insect body, sug-gesting its plausible role in sugar metabolism. The overall understanding of HaST diversity and expression dynamics highlights their putative roles in sugar transport during adaptation and stress recovery of insects.

10aGene expression10aHelicoverpa armigera10aStress10aTransporter-like10atrehalose1 aTellis, Meenakshi, B.1 aChaudhari, Bhagyashri, Y.1 aDeshpande, Shweta, V.1 aNikam, Sanika, V.1 aBarvkar, Vitthal, T.1 aKotkar, Hemlata, M.1 aJoshi, Rakesh, S. uhttps://library.ncl.res.in/content/trehalose-transporter-gene-diversity-and-dynamics-enhances-stress-response-and-recovery02668nas a2200253 4500008004100000022001400041245007600055210006900131260000800200490000600208520183100214653001602045653002602061653001402087653003802101100002602139700002402165700002602189700003002215700001802245700002402263700002202287856010502309 2024 eng d a2701-019800aInhibition of trehalose synthesis in lepidoptera reduces larval fitness0 aInhibition of trehalose synthesis in lepidoptera reduces larval  cFEB0 v83 a

Trehalose is synthesized in insects through the trehalose 6-phosphate synthase and phosphatase (TPS/TPP) pathway. TPP dephosphorylates trehalose 6-phosphate to release trehalose. Trehalose is involved in metamorphosis, but its relation with body weight, size, and developmental timing is unexplored. The expression and activity of TPS/TPP fluctuate depending on trehalose demand. Thus, TPS/TPP inhibition can highlight the significance of trehalose in insect physiology. TPS/TPP transcript levels are elevated in the pre-pupal and pupal stages in Helicoverpa armigera. The inhibition of recombinantly expressed TPP by N-(phenylthio)phthalimide (NPP), is validated by in vitro assays. In vivo inhibition of trehalose synthesis reduces larval weight and size, hampers metamorphosis, and reduces its overall fitness. Insufficient trehalose leads to a shift in glucose flux, reduced energy, and dysregulated fatty acid oxidation. Metabolomics reaffirms the depletion of trehalose, glucose, glucose 6-phosphate, and suppressed tricarboxylic acid cycle. Reduced trehalose hampers the energy level affecting larval vitality. Through trehalose synthesis inhibition, the importance of trehalose in insect physiology and development is investigated. Also, in two other lepidopterans, TPP inhibition impedes physiology and survival. NPP is also found to be effective as an insecticidal formulation. Overall, trehalose levels affect the larval size, weight, and metabolic homeostasis for larval-pupal transition in lepidoptera. Inhibition of trehalose synthesis enzyme TPS/TPP is a good model to study the contribution of trehalose in attaining body weight and in larval-pupal transition. Transcriptomics and metabolomics studies reveal overall reduced energy and dysregulated glucose, protein, and fatty acid metabolism.image

10aLepidoptera10ametabolic homeostasis10atrehalose10atrehalose 6-phosphate phosphatase1 aTellis, Meenakshi, B.1 aMohite, Sharada, D.1 aNair, Vineetkumar, S.1 aChaudhari, Bhagyashri, Y.1 aAhmed, Shadab1 aKotkar, Hemlata, M.1 aJoshi, Rakesh, S. uhttps://library.ncl.res.in/content/inhibition-trehalose-synthesis-lepidoptera-reduces-larval-fitness