00703nas a2200205 4500008004100000022001400041245008000055210006900135260000800204300000900212490000700221653001200228653002100240653003500261100002900296700002200325700002100347700002000368856010900388 2014 eng d a2053-273300aStructural basis of Iridoid synthase mediated cyclization of 10-oxogeranial0 aStructural basis of Iridoid synthase mediated cyclization of 10o cMAY aC2700 v7010aCryo-EM10airidoid synthase10aSingle Particle reconstruction1 aSandholu, Anandsukeerthi1 aThulasiram, H. V.1 aSengupta, Jayati1 aKulkarni, Kiran uhttps://library.ncl.res.in/content/structural-basis-iridoid-synthase-mediated-cyclization-10-oxogeranial00533nas a2200181 4500008004100000022001400041245005400055210005400109260000800163300000900171490000700180653001300187653000900200653001900209100002000228700002000248856008300268 2014 eng d a2053-273300aStructural studies on an atypical Rho GTPase RhoH0 aStructural studies on an atypical Rho GTPase RhoH cMAY aC3890 v7010alymphoma10aRhoH10aTCR signalling1 aMotiwala, Zenia1 aKulkarni, Kiran uhttps://library.ncl.res.in/content/structural-studies-atypical-rho-gtpase-rhoh02366nas a2200277 4500008004100000022001400041245009100055210006900146260009000215300001400305490000800319520140000327653003101727653001501758653002701773653002801800653001401828100002101842700001501863700001701878700002001895700001701915700002101932700001901953856011601972 2015 eng d a0022-283600aAtomic-resolution structures of the APC/C subunits Apc4 and the Apc5 N-terminal domain0 aAtomicresolution structures of the APCC subunits Apc4 and the Ap a24-28 OVAL RD, LONDON NW1 7DX, ENGLANDbACADEMIC PRESS LTD- ELSEVIER SCIENCE LTDcOCT a3300-33150 v4273 a

Many essential biological processes are mediated by complex molecular machines comprising multiple subunits. Knowledge on the architecture of individual subunits and their positions within the overall multimeric complex is key to understanding the molecular mechanisms of macromolecular assemblies. The anaphase-promoting complex/cyclosome (APC/C) is a large multisubunit complex that regulates cell cycle progression by ubiquitinating cell cycle proteins for proteolysis by the proteasome. The holo-complex is composed of 15 different proteins that assemble to generate a complex of 20 subunits. Here, we describe the crystal structures of Apc4 and the N-terminal domain of Apc5 (Apc5(N)). Apc4 comprises a WD40 domain split by a long alpha-helical domain, whereas Apc5(N) has an alpha-helical fold. In a separate study, we had fitted these atomic models to a 3.6-angstrom-resolution cryo-electron microscopy map of the APC/C. We describe how, in the context of the APC/C, regions of Apc4 disordered in the crystal assume order through contacts to Apc5, whereas Apc5(N) shows small conformational changes relative to its crystal structure. We discuss the complementary approaches of high-resolution electron microscopy and protein crystallography to the structure determination of subunits of multimeric complexes. (c) 2015 MRC Laboratory of Molecular Biology. Published by Elsevier Ltd.

10aanaphase-promoting complex10acell cycle10amultisubunit structure10aprotein crystallography10aubiquitin1 aCronin, Nora, B.1 aYang, Jing1 aZhang, Ziguo1 aKulkarni, Kiran1 aChang, Leifu1 aYamano, Hiroyuki1 aBarford, David uhttps://library.ncl.res.in/content/atomic-resolution-structures-apcc-subunits-apc4-and-apc5-n-terminal-domain-001961nas a2200157 4500008004100000245011600041210006900157260000800226300002600234490000600260520135200266100001801618700002301636700002001659856012401679 2017 eng d00aNucleotide dependent switching in Rho GTPase: conformational heterogeneity and competing molecular interactions0 aNucleotide dependent switching in Rho GTPase conformational hete cAPR aArticle Number: 458290 v73 aRas superfamily of GTPases regulate myriad cellular processes through a conserved nucleotide (GTP/ GDP) dependent switching mechanism. Unlike Ras family of GTPases, for the Rho GTPases, there is no clear evidence for the existence of "sub-states" such as state 1 & state 2 in the GTP bound form. To explore the nucleotide dependent conformational space of the Switch I loop and also to look for existence of state 1 like conformations in Rho GTPases, atomistic molecular dynamics and metadynamics simulations on RhoA were performed. These studies demonstrate that both the nucleotide-free state and the GDP bound "OFF" state have very similar conformations, whereas the GTP bound "ON" state has unique conformations with signatures of two intermediate states. The conformational free energy landscape for these systems suggests the presence of multiple intermediate states. Interestingly, the energetic penalty of exposing the non-polar residues in the GTP bound form is counter balanced by the favourable hydrogen bonded interactions between the gamma- phosphate group of GTP with the highly conserved Tyr34 and Thr37 residues. These competing molecular interactions lead to a tuneable energy landscape of the Switch I conformation, which can undergo significant changes based on the local environment including changes upon binding to effectors.1 aKumawat, Amit1 aChakrabarty, Suman1 aKulkarni, Kiran uhttps://library.ncl.res.in/content/nucleotide-dependent-switching-rho-gtpase-conformational-heterogeneity-and-competing01813nas a2200265 4500008004100000022001400041245010000055210006900155260000800224300001400232490000800246520090300254653002101157653001301178653003501191653003401226653002601260100002401286700002001310700002201330700003701352700002001389700002001409856011801429 2018 eng d a1873-346800aDynamics of loops at the substrate entry channel determine the specificity of iridoid synthases0 aDynamics of loops at the substrate entry channel determine the s cAUG a2624-26350 v5923 a

Iridoid synthases belong to the family of short-chain dehydrogenase reductase involved in the biosynthesis of iridoids. Despite having high sequence and structural homology with progesterone 5 beta- reductase, these enzymes exhibit differential substrate specificities. Previously, two loops. L1 and L2 at substrate-binding pocket, were suggested to be involved in generating substrate specificity. However, the structural basis of specificity determinants was elusive. Here, combining sequence and structural analysis, site-directed mutagenesis, and molecular dynamics simulations, we have shown that iridoid synthase contains two channels for substrate entry whose geometries are altered by L1-L2 dynamics, primarily orchestrated by interactions of residues Glu161 and Gly162 of L1 and Asn358 of L2. A complex interplay of these interactions confer the substrate specificity to the enzyme.

10airidoid synthase10airidoids10aMolecular dynamics simulations10aprogesterone 5 beta-reductase10aSubstrate specificity1 aSandholu, Anand, S.1 aMohole, Madhura1 aDuax, William, L.1 aThulasiram, Hirekodathakallu, V.1 aSengupta, Durba1 aKulkarni, Kiran uhttps://library.ncl.res.in/content/dynamics-loops-substrate-entry-channel-determine-specificity-iridoid-synthases02023nas a2200265 4500008004100000022001400041245013300055210006900188260000800257300001400265490000700279520104800286653003701334653002501371653003101396653003301427653001401460653002601474100002401500700002601524700002701550700003701577700002001614856012301634 2020 eng d a0887-358500aStructural studies on 10-hydroxygeraniol dehydrogenase: a novel linear substrate-specific dehydrogenase from Catharanthus roseus0 aStructural studies on 10hydroxygeraniol dehydrogenase a novel li cSEP a1197-12060 v883 a

Conversion of 10-hydroxygeraniol to 10-oxogeranial is a crucial step in iridoid biosynthesis. This reaction is catalyzed by a zinc-dependent alcohol dehydrogenase, 10-hydroxygeraniol dehydrogenase, belonging to the family of medium-chain dehydrogenase/reductase (MDR). Here, we report the crystal structures of a novel 10-hydroxygeraniol dehydrogenase from Catharanthus roseus in its apo and nicotinamide adenine dinucleotide phosphate (NADP(+)) bound forms. Structural analysis and docking studies reveal how subtle conformational differences of loops L1, L2, L3, and helix alpha 9' at the orifice of the catalytic site confer differential activity of the enzyme toward various substrates, by modulating the binding pocket shape and volume. The present study, first of its kind, provides insights into the structural basis of substrate specificity of MDRs specific to linear substrates. Furthermore, comparison of apo and NADP(+) bound structures suggests that the enzyme adopts open and closed states to facilitate cofactor binding.

10a10-hydroxygeraniol dehydrogenase10airidoid biosynthesis10amedium-chain dehydrogenase10amonoterpene indole alkaloids10areductase10aX-ray crystallography1 aSandholu, Anand, S.1 aMujawar, Sharmila, P.1 aRamakrishnan, Krithika1 aThulasiram, Hirekodathakallu, V.1 aKulkarni, Kiran uhttps://library.ncl.res.in/content/structural-studies-10-hydroxygeraniol-dehydrogenase-novel-linear-substrate-specific02242nas a2200241 4500008004100000022001400041245013700055210006900192260000900261300001200270490000800282520138800290653002301678653002401701653002801725653003401753653002601787100001701813700000501830700002001835700002001855856012501875 2021 eng d a0006-291X00aDynamic coupling analysis on plant sesquiterpene synthases provides leads for the identification of product specificity determinants0 aDynamic coupling analysis on plant sesquiterpene synthases provi cJAN  a107-1140 v5363 a

Sesquiterpene synthases catalyse cyclisation of farnesyl pyrophosphate to produce diverse sesquiterpenes. Despite utilising the same substrate and exhibiting significant sequence and structural homology, these enzymes form different products. Previous efforts were based on identifying the effect of divergent residues present at the catalytic binding pocket on the product specificity of these enzymes. However, the rationales deduced for the product specificity from these studies were not generic enough to be applicable to other phylogenetically distant members of this family. To address this problem, we have developed a novel approach combining sequence, structural and dynamical information of plant sesquiterpene synthases (SSQs) to predict product modulating residues (PMRs). We tested this approach on the SSQs with known PMRs and also on sesquisabinene synthase 1 (SaSQS1), a SSQ from Indian sandalwood. Our results show that the dynamical sectors of SSQs obtained from molecular dynamics simulation and their hydrophobicity and vicinity indices together provide leads for the identification of PMRs. The efficacy of the technique was tested on SaSQS1 using mutagenesis. To the best of our knowledge, this is a first technique of this kind which provides cues on PMRs of SSQs, with divergent phylogenetic relationship. (C) 2020 Elsevier Inc. All rights reserved.

10amolecular dynamics10aProduct specificity10aSesquiterpene synthases10aStatistical coupling analysis10aX-ray crystallography1 aSingh, Sneha1 a1 aSengupta, Durba1 aKulkarni, Kiran uhttps://library.ncl.res.in/content/dynamic-coupling-analysis-plant-sesquiterpene-synthases-provides-leads-identification01988nas a2200229 4500008004100000022001400041245009800055210006900153260000800222300001000230490000800240520120800248653001701456653003701473653001801510100002001528700001701548700002101565700002601586700002001612856012601632 2021 eng d a0022-263100aExpression, purification and crystallization of Asrij, A novel scaffold transmembrane protein0 aExpression purification and crystallization of Asrij A novel sca cFEB a65-740 v2543 a

Asrij/OCIAD1 is a scaffold transmembrane protein belonging to the Ovarian Carcinoma Immunoreactive Antigen Domain containing protein family. In Drosophila and mouse models, Asrij localizes at the endosomal and mitochondrial membrane and is shown to regulate the stemness of hematopoietic stem cells. Interaction of Asrij with ADP Ribosylation Factor 1 (Arf1) is shown to be crucial for hematopoietic niche function and prohemocyte maintenance. Here, we report the heterologous expression, standardization of detergents and purification methodologies for crystallization of Asrij/OCIAD1. To probe the activity of bacterially expressed Asrij, we developed a protein complementation assay and conclusively show that Asrij and Arf1 physically interact. Further, we find that sophorolipids improve the solubility and monodispersibility of Asrij. Hence, we propose that sophorolipids could be novel additives for stabilization of membrane proteins. To our knowledge, this is the first study detailing methodology for the production and crystallization of a heterologously expressed scaffold membrane protein and will be widely applicable to understand membrane protein structure and function. Graphic

10aAsrij/OCIAD110aMembrane protein crystallography10asophorolipids1 aMotiwala, Zenia1 aDarne, Priti1 aPrabhune, Asmita1 aInamdar, Maneesha, S.1 aKulkarni, Kiran uhttps://library.ncl.res.in/content/expression-purification-and-crystallization-asrij-novel-scaffold-transmembrane-protein01809nas a2200133 4500008004100000245006200041210006200103260000800165300000900173520136400182100002101546700002001567856008801587 2021 eng d00aGPCRs that Rh oar the Guanine nucleotide exchange factors0 aGPCRs that Rh oar the Guanine nucleotide exchange factors cAPR a1-163 a

Cell migration, a crucial step in numerous biological processes, is tightly regulated in space and time. Cells employ Rho GTPases, primarily Rho, Rac, and Cdc42, to regulate their motility. Like other small G proteins, Rho GTPases function as biomolecular switches in regulating cell migration by operating between GDP bound 'OFF' and GTP bound 'ON' states. Guanine nucleotide exchange factors (GEFs) catalyse the shuttling of GTPases from OFF to ON state. G protein-coupled receptors (GPCRs) are the largest family of cell surface receptors that are involved in many signalling phenomena including cell survival and cell migration events. In this review, we summarize signalling mechanisms, involving GPCRs, leading to the activation of RhoGEFs. GPCRs exhibit diverse GEF activation modes that include the interaction of heterotrimeric G protein subunits with different domains of GEFs, phosphorylation, protein-protein interaction, protein-lipid interaction, and/or a combination of these processes.

1 aOmble, Aishwarya1 aKulkarni, Kiran uhttps://library.ncl.res.in/content/gpcrs-rh-oar-guanine-nucleotide-exchange-factors01754nas a2200241 4500008004100000022001400041245010500055210006900160260000900229300001100238490000700249520094300256653001601199653002101215653001801236653002501254653002201279100002101301700002601322700002001348700002401368856012001392 2021 eng d a0960-894X00aMachine learning classifiers aid virtual screening for efficient design of mini-protein therapeutics0 aMachine learning classifiers aid virtual screening for efficient cAPR  a1278520 v383 a

De novo design of mini-proteins (4-12 kDa) has recently been shown to produce new candidates for protein therapeutics. They are temperature stable molecules that bind to the drug target with high affinity for inhibiting its interactions. The development of mini-protein binders requires laboratory screening of tens of thousands of molecules for effective target binding. In this study we trained machine learning classifiers which can distinguish, with 90% accuracy and 80% precision, mini-protein binders from non-binding molecules designed for a particular target; this significantly reduces the number of mini protein candidates for experimental screening. Further, on the basis of our results we propose a multi-stage protocol where a small dataset (few hundred experimentally verified target-specific mini-proteins) can be used to train classifiers for improving the efficiency of mini-protein design for any specific target.

10aDrug design10amachine learning10aMini-proteins10aProtein therapeutics10avirtual screening1 aGaur, Neeraj, K.1 aGoyal, Venuka, Durani1 aKulkarni, Kiran1 aMakde, Ravindra, D. uhttps://library.ncl.res.in/content/machine-learning-classifiers-aid-virtual-screening-efficient-design-mini-protein00544nas a2200157 4500008004100000022001400041245007400055210006900129260000800198300001200206490000800218100002100226700002000247700002000267856009900287 2021 eng d a0022-263100aMolecular interplay at the membrane and impact on cellular physiology0 aMolecular interplay at the membrane and impact on cellular physi cJUN a239-2420 v2541 aPucadyil, Thomas1 aKulkarni, Kiran1 aSengupta, Durba uhttps://library.ncl.res.in/content/molecular-interplay-membrane-and-impact-cellular-physiology02873nas a2200241 4500008004100000022001400041245014800055210006900203260000800272300001000280490000800290520199200298653002902290653002302319653002902342653003502371653002302406653001602429100001902445700002002464700002002484856012702504 2022 eng d a0006-291X00aSubstrate induced dynamical remodeling of the binding pocket generates GTPase specificity in DOCK family of guanine nucleotide exchange factors0 aSubstrate induced dynamical remodeling of the binding pocket gen cNOV a32-400 v6313 a

Dedicator of cytokinesis (DOCK) family of guanine nucleotide exchange factors (GEFs) activate two members of Rho family GTPases, Rac1/Cdc42, to exert diverse cellular processes, including cell migration. As DOCK GEFs have been critically implicated in tumour cell migration, understanding their function and specificity is imperative for designing anti-metastatic drugs. Based on their GTPase specificity they have been classified as Rac, Cdc42 and dual specific GEFs. Despite extensive structural studies, the factors that determine GTPase specificity of DOCK GEFs have remained elusive. Here, we show that subtle dynamical coupling between GEF and GTPase structures modulate the binding interface to generate mutual spec-ificity. To cluster the dynamically coupled residues in GEF-GTPase complexes a novel intra-residue backbone-torsion-angles based mutual information (TMI) technique was employed. TMI was calcu-lated from 4500 trajectories obtained from a total of 4.5ms molecular dynamics simulations performed on members of all the three clades of DOCK GEFs. The obtained clusters suggest a specificity generation mechanism that involves optimization of the binding pocket for the crucial divergent residue at the 56th position of Rac/Cdc42 (FCdc42/WRac1). These clusters encompass five residues from the structural segment lobe C -a10 helix of the DOCK proteins and functional SWI region of GTPase, which induce orchestrated structural modulations to generate the specificity. Even the conserved residues from SWI region are seen to augment the specificity defining dynamical rearrangements. Furthermore, the pro-posed dynamical GTPase-DOCK GEF specificity model was verified using mutagenesis studies on Rac1 and dual GTPase specific Dock2 and Dock6, respectively. Thus the current study provides the generic substrate specificity determinants of DOCK GEFs, which are not apparent from the conventional struc-tural analysis.(c) 2022 Elsevier Inc. All rights reserved.

10aDedicator of cytokinesis10aGTPase specificity10aGuanine exchange factors10aMolecular dynamics simulations10aMutual information10aRho GTPases1 aRoy, Debopriya1 aSengupta, Durba1 aKulkarni, Kiran uhttps://library.ncl.res.in/content/substrate-induced-dynamical-remodeling-binding-pocket-generates-gtpase-specificity-dock03041nas a2200193 4500008004100000245013000041210006900171260000900240300001400249490000700263520232300270100001902593700002202612700001902634700002002653700002002673700002402693856013002717 2023 eng d00aCocrystallization of multi-kinase inhibitor pazopanib with fenamic acids: improving dissolution and inhibiting cell migration0 aCocrystallization of multikinase inhibitor pazopanib with fenami cOCT  a5565-55740 v253 a

A multi-kinase inhibitor, pazopanib (PAZ) is cocrystallized with cyclooxygenase (COX) inhibitor fenamic acids to investigate the dissolution rate and inhibition of cell migration in VEGF-triggered HUVEC cells to test the efficacy of stoichiometric drug-drug combinations. Crystallization experiments at the sub-milligram level in an acetonitrile-methanol mixture yielded two drug-drug salt forms of PAZ with flufenamic acid (FFA) and niflumic acid (NFA) as PAZ+center dot FFA-center dot ACN (an acetonitrile solvate named as form I) and PAZ+center dot NFA-. Structures of the crystal forms were characterized by single crystal X-ray diffraction (SC-XRD) method. Crystal structures revealed that the presence of a 2-aminopyrimidine group in PAZ is a strong partner for the carboxyl group in all solid forms, forming an acidMIDLINE HORIZONTAL ELLIPSISpyrimidine heterosynthon with COX inhibitor fenamic acids. To perform dissolution experiments and cell line analysis, the scale-up of both salt forms were done in the acetonitrile-methanol mixture through crystallization, which showed a polymorphic transformation in the case of PAZ+center dot FFA-center dot ACN (an acetonitrile solvate named form II). The thermodynamic stability of PAZ+center dot FFA-center dot ACN (form II) and PAZ+center dot NFA- were analysed using slurry experiment under ambient conditions in pH 1.2 (0.1 N HCl) buffer medium and the residual solid phase was characterized by powder XRD, which showed that PAZ+center dot FFA-center dot ACN (form II) was a metastable solid form while PAZ+center dot NFA- was a stable solid form. The dissolution experiments at gastric pH 1.2 showed that the rate of dissolution of PAZ+center dot FFA-center dot ACN was 10 times higher than PAZ+center dot NFA-. The cell migration assay suggested that PAZ+center dot FFA-center dot ACN inhibited similar to 25% and PAZ+center dot NFA- inhibited similar to 20% migration of HUVEC cells compared to PAZ alone. These investigations suggested that the drug-drug salts PAZ+center dot FFA- and PAZ+center dot NFA- would be potential combo drug candidates for clinical trials. Multi-drug crystalline salts of pazopanib with fenamic acids exhibit enhanced dissolution and 20% higher inhibition in the migration of HUVEC cells compared to the reference drug.

1 aRai, Sunil, K.1 aGunnam, Anilkumar1 aRoy, Debopriya1 aRajput, Raveena1 aKulkarni, Kiran1 aNangia, Ashwini, K. uhttps://library.ncl.res.in/content/cocrystallization-multi-kinase-inhibitor-pazopanib-fenamic-acids-improving-dissolution-and02551nas a2200253 4500008004100000022001400041245013600055210006900191260000900260300001400269490000700283520166300290653002601953653001701979653002001996653002302016653001802039100002102057700001802078700002602096700002002122700002402142856013102166 2023 eng d a0739-110200aEvolutionary conservation of protein dynamics: insights from all-atom molecular dynamics simulations of `peptidase' domain of Spt160 aEvolutionary conservation of protein dynamics insights from alla cMAR  a1445-14570 v413 a

Protein function is encoded in its sequence, manifested in its three-dimensional structure, and facilitated by its dynamics. Studies have suggested that protein structures with higher sequence similarity could have more similar patterns of dynamics. However, such studies of protein dynamics within and across protein families typically rely on coarse-grained models, or approximate metrics like crystallographic B-factors. This study uses mu s scale molecular dynamics (MD) simulations to explore the conservation of dynamics among homologs of similar to 50 kDa N-terminal module of Spt16 (Spt16N). Spt16N from Saccharomyces cerevisiae (Sc-Spt16N) and three of its homologs with 30-40% sequence identities were available in the PDB. To make our data-set more comprehensive, the crystal structure of an additional homolog (62% sequence identity with Sc-Spt16N) was solved at 1.7 angstrom resolution. Cumulative MD simulations of 6 mu s were carried out on these Spt16N structures and on two additional protein structures with varying degrees of similarity to it. The simulations revealed that correlation in patterns of backbone fluctuations vary linearly with sequence identity. This trend could not be inferred using crystallographic B-factors. Further, normal mode analysis suggested a similar pattern of inter-domain (inter-lobe) motions not only among Spt16N homologs, but also in the M24 peptidase structure. On the other hand, MD simulation results highlighted conserved motions that were found unique for Spt16N protein, this along with electrostatics trends shed light on functional aspects of Spt16N. Communicated by Ramaswamy H. Sarma.

10aBackbone fluctuations10aFACT complex10ahistone binding10ainterdomain motion10aM24 peptidase1 aGaur, Neeraj, K.1 aGhosh, Biplab1 aGoyal, Venuka, Durani1 aKulkarni, Kiran1 aMakde, Ravindra, D. uhttps://library.ncl.res.in/content/evolutionary-conservation-protein-dynamics-insights-all-atom-molecular-dynamics-simulations03066nas a2200277 4500008004100000022001400041245013100055210006900186260000800255300001200263490000700275520214700282653000902429653002302438653001802461653001002479653001402489653003502503100001702538700001702555700001802572700002502590700002002615700002602635856012702661 2023 eng d a2296-634X00aIn silico identification and characterization of the SNPs in the human ASTL gene and their probable role in female infertility0 aIn silico identification and characterization of the SNPs in the cJUN a11516720 v113 a

Ovastacin (ASTL), a zinc metalloprotease, is released from a fertilized egg during exocytosis of cortical granules which occurs minutes after the sperm and egg fuse. ASTL cleaves ZP2, one of the four primary glycoproteins of human zona pellucida, and this cleavage prevents polyspermy, causes zona pellucida hardening, and also protects the pre-implantation embryo. Any perturbation in the activity of ASTL can thus disturb this process and may lead to infertility without changing the gross morphology of the oocyte. A small amount of ASTL is also released by unfertilized oocytes but its catalytic activity is absent as it is bound by its inhibitor, Fetuin-B (FETUB). Pre-mature release of ASTL when FETUB is absent also causes infertility. To identify and understand the structural and functional effects of deleterious SNPs of ASTL on its interaction with ZP2 and FETUB and hence on fertility, a total of 4,748 SNPs from the dbSNP database were evaluated using a variety of in silico tools. All of the 40 shortlisted nsSNPs were present in the catalytic domain of the protein. Comparison of the wild type with mutants using MutPred2 suggests an alteration in the catalytic activity/zinc binding site in many SNPs. Docking studies show the involvement of hydrophobic interactions and H bonding between ASTL and ZP2 and also between ASTL and FETUB. Four positions in ASTL involved in the hydrophobic interactions (P-105 and D-200 between ASTL and ZP2; D-198 and L-278 etween ASTL and FETUB) and 5 in H bonding (E-75 and R-159 between ASTL and ZP2; and K-93, R-159, and C-281 between ASTL and FETUB) have SNP's associated with them validating their importance. Interestingly, a cluster of multiple SNPs was found in the motif (DRD200)-D-198, which is also a well-conserved region among several species. Statistical Coupling Analysis (SCA) suggested that the deleterious SNPs were present in the functionally important amino acid positions of ASTL and are evolutionarily coupled. Thus, these results attempt to identify the regions in ASTL, mutations in which can affect its binding with ZP2 or FETUB and cause female infertility.

10aASTL10afemale infertility10afertilization10aomics10aovastacin10asingle nucleotide polymorphism1 aSuri, Kapali1 aRajput, Neha1 aSharma, Priya1 aOmble, Aishwarya, D.1 aKulkarni, Kiran1 aGahlay, Gagandeep, K. uhttps://library.ncl.res.in/content/silico-identification-and-characterization-snps-human-astl-gene-and-their-probable-role01658nas a2200169 4500008004100000245010500041210006900146260000800215300001100223490000700234520104200241100002001283700002401303700002001327700002001347856012101367 2023 eng d00aWavelet coherence phase analysis decodes the universal switching mechanism of Ras GTPase superfamily0 aWavelet coherence phase analysis decodes the universal switching cJUL a1070310 v263 a

The Ras superfamily of GTPases regulate critical cellular processes by shuttling between GTP-bound ON and GDP-bound OFF states. This switching mechanism is attributed to the conformational changes in two loops, SWI and SWII, upon GTP binding and hydrolysis. Since these conformational changes vary across the Ras superfamily, there is no generic parameter to define their functional states. A unique wavelet coherence (WC) analysis-based approach developed here shows that the structural changes in switch regions could be mapped onto the wavelet coherence phase couplings (WPCs). Thus, WPCs could serve as unique parameters to define their functional states. Disentanglement of WPCs in oncogenic GTPases shows how breakdown of structural allostery leads to their aberrant function. These observations stand out even for simulated ensemble of switch region conformers. Overall, for the first time, we show that WPCs could unravel the latent structural deviations in Ras proteins to decode their universal switching mechanism.

1 aMotiwala, Zenia1 aSandholu, Anand, S.1 aSengupta, Durba1 aKulkarni, Kiran uhttps://library.ncl.res.in/content/wavelet-coherence-phase-analysis-decodes-universal-switching-mechanism-ras-gtpase00738nas a2200205 4500008004100000022001400041245010300055210006900158260000900227300001100236490000800247653001700255100002600272700002400298700002000322700002000342700002400362700002000386856012600406 2024 eng d a0006-291X00aCell based assay using virus-like particles to screen AM type mimics for SARS-CoV-2 neutralisation0 aCell based assay using viruslike particles to screen AM type mim cJUL  a1500820 v71810aMINIPROTEINS1 aGaur, Neeraj, Kailash1 aUrankar, Shreegauri1 aSengupta, Durba1 aChepuri, Ramana1 aMakde, Ravindra, D.1 aKulkarni, Kiran uhttps://library.ncl.res.in/content/cell-based-assay-using-virus-particles-screen-am-type-mimics-sars-cov-2-neutralisation02256nas a2200169 4500008004100000245010000041210006900141260000900210300000800219490000600227520165300233100002101886700002201907700002001929700002001949856011701969 2024 eng d00aDishevelled2 activates WGEF via its interaction with a unique internal peptide motif of the GEF0 aDishevelled2 activates WGEF via its interaction with a unique in cMAY  a5430 v73 a

The Wnt-planar cell polarity (Wnt-PCP) pathway is crucial in establishing cell polarity during development and tissue homoeostasis. This pathway is found to be dysregulated in many pathological conditions, including cancer and autoimmune disorders. The central event in Wnt-PCP pathway is the activation of Weak-similarity guanine nucleotide exchange factor (WGEF) by the adapter protein Dishevelled (Dvl). The PDZ domain of Dishevelled2 (Dvl2PDZ) binds and activates WGEF by releasing it from its autoinhibitory state. However, the actual Dvl2PDZ binding site of WGEF and the consequent activation mechanism of the GEF have remained elusive. Using biochemical and molecular dynamics studies, we show that a unique ``internal-PDZ binding motif'' (IPM) of WGEF mediates the WGEF-Dvl2PDZ interaction to activate the GEF. The residues at P 2 , P 0 , P -2 and P -3 positions of IPM play an important role in stabilizing the WGEFpep-Dvl2PDZ interaction. Furthermore, MD simulations of modelled Dvl2PDZ-WGEFIPM peptide complexes suggest that WGEF-Dvl2PDZ interaction may differ from the reported Dvl2PDZ-IPM interactions. Additionally, the apo structure of human Dvl2PDZ shows conformational dynamics different from its IPM peptide bound state, suggesting an induced fit mechanism for the Dvl2PDZ-peptide interaction. The current study provides a model for Dvl2 induced activation of WGEF. This study identifies a unique ``internal-PDZ binding motif'' (IPM) of WGEF that mediates its interaction with Dvl2PDZ, leading to the release of GEF from its autoinhibitory state. It further unravels the underlying mutual Dvl2-IPM recognition aspects.

1 aOmble, Aishwarya1 aMahajan, Shrutika1 aBhoite, Ashwini1 aKulkarni, Kiran uhttps://library.ncl.res.in/content/dishevelled2-activates-wgef-its-interaction-unique-internal-peptide-motif-gef02559nas a2200265 4500008004100000022001400041245014200055210006900197260000900266300001100275490000800286520163300294653002301927653002201950653003401972653002102006653002602027100002002053700002102073700001902094700002202113700001702135700002002152856012102172 2024 eng d a0006-291X00aStructure of epoxide hydrolase 2 from Mangifera indica throws light on the substrate specificity determinants of plant epoxide hydrolases0 aStructure of epoxide hydrolase 2 from Mangifera indica throws li cNOV  a1504440 v7333 a

Epoxide hydrolases (EHs) are a group of ubiquitous enzymes that catalyze hydrolysis of chemically reactive epoxides to yield corresponding dihydrodiols. Despite extensive studies on EHs from different clades, generic rules governing their substrate specificity determinants have remained elusive. Here, we present structural, biochemical and molecular dynamics simulation studies on MiEH2, a plant epoxide hydrolase from Mangifera indica. Comparative structure-function analysis of nine homologs of MiEH2, which include a few AlphaFold structural models, show that the two conserved tyrosines (MiEH2Y152 and MiEH2Y232) from the lid domain dissect substrate binding tunnel into two halves, forming substrate-binding-pocket one (BP1) and two (BP2). This compartmentalization offers diverse binding modes to their substrates, as exemplified by the binding of smaller aromatic substrates, such as styrene oxide (SO). Docking and molecular dynamics simulations reveal that the linear epoxy fatty acid substrates predominantly occupy BP1, while the aromatic substrates can bind to either BP1 or BP2. Furthermore, SO preferentially binds to BP2, by stacking against catalytically important histidine (MiEH2H297) with the conserved lid tyrosines engaging its epoxide oxygen. Residue (MiEH2L263) next to the catalytic aspartate (MiEH2D262) modulates substrate binding modes. Thus, the divergent binding modes correlate with the differential affinities of the EHs for their substrates. Furthermore, long-range dynamical coupling between the lid and core domains critically influences substrate enantioselectivity in plant EHs.

10aEnantioselectivity10aEpoxide hydrolase10amolecular dynamics simulation10aRegioselectivity10aX-ray crystallography1 aBhoite, Ashwini1 aGaur, Neeraj, K.1 aPalange, Megha1 aKontham, Ravindar1 aGupta, Vidya1 aKulkarni, Kiran uhttps://library.ncl.res.in/content/structure-epoxide-hydrolase-2-mangifera-indica-throws-light-substrate-specificity