Exploring Nocardia's ecological spectrum and novel therapeutic frontiers through whole-genome sequencing: unraveling drug resistance and virulence factors.

Nocardia farcinica is the leading pathogen responsible for nocardiosis, a life-threatening infection primarily affecting immunocompromised patients. In this study, the genomic sequence of a clinically isolated N. farcinica sample was sequenced. Subsequently, the assembled genome was annotated to identify antimicrobial resistance and virulence genes, as well as plasmid and prophages. The analysis of the entire genome size was 6,021,225 bp, with a GC content of 70.78% and consists of 103 contigs and N50 values of 292,531 bp. The genome analysis revealed the presence of several antimicrobial resistance genes, including RbpA, mtrA, FAR-1, blaFAR-1, blaFAR-1_1, and rox. In addition, virulence genes such as relA, icl, and mbtH were also detected. The present study signifies that N. farcinica genome is pivotal for the understanding of antimicrobial resistance and virulence genes is crucial for comprehending resistance mechanism, and developing effective strategies to combat bacterial infections effectively, especially adhesins and toxins. This study aids in identifying crucial drug targets for combating multidrug-resistant N. farcinica in the future.

Repurposing FDA-approved drugs for combating tigecycline resistance in Acinetobacter baumannii: in silico screening against BaeR protein.

Acinetobacter baumannii is becoming a gravely threatening nosocomial infection with a higher mortality rate. The present study targets the BaeR protein that mediates resistance to tigecycline antibiotics. The BaeR protein, along with the aid of BaeS, senses the incoming antibiotics and stimulates the expression of resistance proteins. These resistance proteins efflux the antibiotics and protect the cells from its effect. The main goal of the current study is to determine potential inhibitors from already existing FDA-approved drugs that could mitigate the BaeR protein. A range of in silico approaches, including molecular dynamics, virtual screening, SIFT analysis, ADMET, DFT, MM/GBSA, MMPBSA and per residue interaction analysis, were performed to identify inhibitors against this protein. The screening of FDA-approved compounds against the BaeR protein yielded 620 compounds. These compounds were clustered by SIFT to distinguish related compounds, it resulted in 20 different clusters. The top five clusters that can accommodate the binding site with better interaction and score by fulfilling all criteria were selected. The DFT analysis showed a smaller energy gap among all the compounds, indicating the ability of the compound to form firm interactions. All the compounds showed less binding free energy in both MM/GBSA and MM/PBSA analyses. The compounds were observed to be stable throughout the simulation. The per-residue interaction analysis confirmed that interactions with binding site residues were stable throughout the simulation. As a result of the study, four compounds, namely ZINC000003801919, DB01203, DB11217 and ZINC0000000056652, were identified as efficient candidates to deal with antimicrobial resistance in A. baumannii.

Whole-genome sequencing unravels novel genetic determinants and regulatory pathways associated with triamcinolone acetonide-induced ocular hypertension.

Glucocorticosteroids commonly used to treat certain ocular inflammatory conditions cause an unwarranted elevation in intraocular pressure (IOP) leading to steroid-induced ocular hypertension (OHT). This study aims to identify novel genetic variants in the Indian population associated with steroid responsiveness, specifically to that of intravitreal Triamcinolone acetonide (TA) injections, which leads to OHT in 27% of the TA-treated Indian subjects. Genetic determinants and pathways regulating TA-OHT progression were investigated by applying whole-genome sequencing (WGS) on DNA extracted from 53 blood samples that included TA responders and non-responders. Sequencing analysis yielded 45 intronic and 49 exonic variants to be associated with TA-OHT, which are known to play a vital role in eye, heart, brain, and bone deformities. Of these, the most significant genetic variant associated with TA-OHT was further considered for molecular dynamics (MD) simulation studies. Variants in the CRPPA, PLOD1, ARHGAP1, TIMELESS and TNFSF4 genes were found to be directly implicating TA-OHT. Furthermore, these genes were enriched in pathways associated with cardiomyopathy, focal adhesion, extracellular matrix, and actin cytoskeleton reorganization. MD simulation studies revealed that the top significant variant (rs141625803) in the CRPPA gene possesses a high pathogenic and structurally destabilizing effect. Thus, novel genetic variants that could be significantly associated with the TA-OHT progression were identified in this study. Validation of these targets in a larger cohort of patients along with their functional analysis would inform on the disease, thereby adding to the existing knowledge on the pathophysiology of TA-OHT.

Deciphering the conformational transitions of LIMK2 active and inactive states to ponder specific druggable states through microsecond scale molecular dynamics simulation.

LIMK2 inhibitors are one of the potential therapeutic modalities for treating various diseases. In the current scenario, there is a paucity of effective LIMK inhibitors that are highly specific with minimal off-target effects. To date, the conformational transitions of LIMK2 from DFGinαCin (CIDI) (active) to DFGoutαCout (CODO) (inactive) states are yet to be probed and are essential for capturing the unique, druggable conformations. Therefore, this study was intended to capture the diverse conformational states of LIMK2 for accelerating the rational identification of conformation specific inhibitors through high-end structural bioinformatics protocols. Hence, in this study, molecular modelling followed by an extensive microsecond timescale of molecular dynamics simulation was performed encompassing perturbation response scanning, metapath, and community analysis towards the conformational sampling of LIMK2. Overall this study precisely identifies the conformational ensemble of LIMK2 the intermediate inactive states namely, CIDO, CinterDinter, CIDinter, CinterDI, CinterDO, CODI, CODinter apart from CIDI and CODO. This also facilitated observing that ß8 preceding XDFG, αC (F373, L374), and αD (L413) as the major effectors that may facilitate the regulation of varying conformational transitions among the states. Additionally, the conserved ß sheets and the loops namely, C.l, b.l, and G/P.loop were observed to be involved in the metapath for allosteric communication among the intermediates with CIDI and CODO state. Moreover, only the CODO state was observed to have closed type A.l, while the CIDI and other intermediate states except for CIDO were observed to have open-DFG out type A.l, thereby enabling the binding of substrate. Apart from these, the druggable site analysis inferred that the CIDI and CODO states harbor prominent druggable sites spanning the conserved N-lobe, while the intermediates were observed to have unraveled allosteric druggable sites distal from the ATP binding site, majorly spanning the C-lobe of LIMK2. Thus, this study provides potential insights into the intermediate conformational druggable states of LIMK2 and also the druggable conformations, especially the inactive states of LIMK2, as a specific therapeutic targeting mode. Thus, providing a widened avenue to ponder the allosteric sites or the isoform selectivity conformations for targeting LIMK2 in various disease conditions.

Design of inhibitory peptide targeting Toxoplasma gondii RON4-human ß-tubulin interactions by implementing structural bioinformatics methods.

Toxoplasma gondii an obligate intracellular parasite causes toxoplasmosis in homeothermic animals. Host invasion of this parasite is mediated by the formation of Moving Junction (MJ) complex which encompasses a network of microneme and Rhoptry Neck proteins (RONs) 2/4/5/8. Among these proteins, RON4 is the only cytosolic secretory protein that is considered as a crucial member, as it directly facilitates the motility of MJ complex by interacting with host tubulin. It is also prominently localized at the host-pathogen interface during the invasion, thus projecting it as a potential drug target. The structure of RON4 is yet to be crystallized. Hence, in this study, fold recognition and Free Energy Landscape sampling was performed to predict the plausible 3D structure of RON4. Further, its interacting pattern with the reported crystal structure of human tubulin was analyzed using molecular docking. Subsequently, a ß-tubulin based inhibitory peptides were derived based on its interacting interface observed in RON4-ß-tubulin docked complex. Following which, a stepwise validation of these peptides for various physico-chemical properties and its homology with antimicrobial peptides were also screened. The peptide (RT_pep) surpassing all these validation filters was modeled and its stability was analysed by Molecular Dynamics simulation. To validate further, the stable conformation of the RT_pep was docked to RON4. Finally, essential molecular dynamics simulation was conducted to determine the stability and atomic motions of native RON4 and also to decipher its association with ß-tubulin and RT_pep. All these analyses cumulatively suggest the therapeutic potential of RT_pep in targeting toxoplasmosis.

Demystifying the pH dependent conformational changes of human heparanase pertaining to structure-function relationships: an in silico approach.

Heparanase (HPSE) is an endo-ß-D-glucuronidase that has diverse functions in mammals which includes cell survival, cell adhesion and cell migration. HPSE features both enzymatic and non-enzymatic functionalities in a pH dependent manner. Hence, in this study, an extensive molecular dynamics simulation, molecular docking, protein Angular dispersion analysis were performed for apo form and holo forms to understand its conformational changes at varied pH conditions. On comparative conformational analysis of apo and holo forms, it was inferred that the HSPE has undergone pH dependent structural changes, thereby affecting the binding of Heparan sulfate proteoglycan (HSPG). Moreover, HPSE also showed favourable structural changes for optimal binding of HSPG at pH 5.0 and 6.0, as inferred from functional flap displacements within HPSE. Thus, this study provides significant insights on optimal pH for HPSE to exhibit its enzymatic activity. The outcome of this study shall aid in ideal lead generation for targeting HPSE mediated disease conditions.

Human Antigen R -mediated modulation of Transforming Growth Factor Beta 1 expression in retinal pathological milieu.

The fate and stability of messenger RNA (mRNA), from transcription to degradation is regulated by a dynamic shuttle of epigenetic modifications and RNA binding proteins in maintaining healthy cellular homeostasis and disease development. While Transforming Growth Factor Beta 1 (TGFß1) has been implicated as a key regulator for diabetic retinopathy, a microvascular complication of diabetes, the RNA binding proteins post-transcriptionally regulating its expression remain unreported in the ocular context. Further, dysfunction of TGFß1 signalling is also strongly associated with angiogenesis, inflammatory responses and tissue fibrosis in many eye conditions leading to vision loss. In this study, computational and molecular simulations were initially carried out to identify Human Antigen R (HuR) binding sites in TGFß1 mRNA and predict the structural stability of these RNA-protein interactions. These findings were further validated through in vitro experiments utilizing Cobalt Chloride (CoCl2) as a hypoxia mimetic agent in human retinal microvascular endothelial cells (HRMVEC). In silico analysis revealed that HuR preferentially binds to the 5'-UTR of TGFß1 and displayed more stable interaction than the 3'UTR. Consistent with in silico analysis, RNA immunoprecipitation demonstrated a robust association between HuR and TGFß1 mRNA specifically under hypoxic conditions. Further, silencing of HuR significantly reduced TGFß1 protein expression upon CoCl2 treatment. Thus, for the first time in ocular pathological milieu, direct evidence of HuR- TGFß1 mRNA interaction under conditions of hypoxia has been reported in this study providing valuable insights into RNA binding proteins as therapeutic targets for ocular diseases associated with TGFß1 dysregulation.

Biochemical and biophysical characterization of biosynthetic arginine decarboxylase from Thermus thermophilus.

The biosynthetic arginine decarboxylase in Thermus thermophilus is responsible for producing spermidine, a polyamine with numerous biological applications in humans. The arginine decarboxylase has significant applications in biotechnology industries, suggesting the need to evaluate its biochemical and biophysical characteristics at the molecular level. In this study, both in vitro and in silico methods were employed to investigate the structural and functional behavior of the arginine decarboxylase protein. In in vitro, MALDI-TOF, size exclusion, and assay studies were performed to examine the nature and activity of the protein. The MALDI-TOF analysis confirmed the purified protein as biosynthetic arginine decarboxylase. The assay results revealed that the Pyridoxal 5'-Phosphate (PLP) cofactor plays a crucial role in enhancing enzyme activity by producing agmatine (a by-product of spermidine). Further, optimum enzyme activity was observed at 50 °C, suggesting the extremophilic nature of the enzyme. Unlike other proteins, this enzyme displayed optimal activity at both acidic and basic pH, demonstrating its sensitivity to pH changes. Furthermore, the addition of divalent ions like Mg 2+ increased the rate of reaction. In in silico, structure modeling, and comparative molecular dynamics simulation studies were used to assess the protein stability and behavior at different pH and temperature conditions. The findings of this study could be applied to improve enzyme production in the industry.Communicated by Ramaswamy H. Sarma.

Unveiling the therapeutic potential of a mutated paraoxonase 2 in diabetic retinopathy: Defying glycation, mitigating oxidative stress, ER stress and inflammation.

Paraoxonase 2 (PON2) is an intracellular anti-oxidant protein ubiquitously expressed in all cells and reduces reactive oxygen species, endoplasmic reticulum (ER) stress, further improves mitochondrial function and thereby shows anti-apoptotic function. In diabetes and its complications this PON gets glycated and becomes in effective. The PON activity is reported to be reduced in diabetic retinopathy and we have earlier showed Carboxy methyl lysine (AGE) decreased PON2 expression and activity in Human retinal endothelial cells (HREC) . In this study, we have designed and developed a mutated PON2 by in silico and in vitro approach which can resist glycation. Where in glycation-prone residues in PON2 was predicted using in silico analyses and a mutated PON2 was developed using in vitro site directed mutagenesis (SDM) assay mPON2 (mutant PON2-PON2-K70A) and its efficacy was compared with wPON2 (wild type PON2). CML glycated wPON2 and reduced its activity when compared with mPON2 in HREC confirmed by immunoprecipitation and in vitro experiments. Additionally, mPON2 interaction efficiency with its substrates was higher than wPON2 by insilico assay and demonstrated enhanced inhibition against CML-induced oxidative stress, ER stress, pro-inflammation, and mitochondrial fission than wPON2 by invitro assay. Further mPON2 showed increased inhibition of phosphorylation of NFĸB induced by CML. Our investigation establishes that the over expression of mPON2 in HREC can defy glycation and therefore mitigate ER stress and inflammation against CML than endogenous wPON2. These findings imply that mPON2 can be a beneficial therapeutic target against diabetic retinopathy.

Deciphering the pH-dependent oligomerization of aspartate semialdehyde dehydrogenase from Wolbachia endosymbiont of Brugia malayi: An in vitro and in silico approaches.

The enzyme aspartate semialdehyde dehydrogenase (ASDH) plays a pivotal role in the amino acid biosynthesis pathway, making it an attractive target for the development of new antimicrobial drugs due to its absence in humans. This study aims to investigate the presence of ASDH in the filarial parasite Wolbachia endosymbiont of Brugia malayi (WBm) using both in vitro and in silico approaches. The size exclusion chromatography (SEC) and Native-PAGE analysis demonstrate that WBm-ASDH undergoes pH-dependent oligomerization and dimerization. To gain a deeper understanding of this phenomenon, the modelled monomer and dimer structures were subjected to pH-dependent dynamics simulations in various conditions. The results reveal that residues Val240, Gln161, Thr159, Tyr160, and Trp316 form strong hydrogen bond contacts in the intersurface area to maintain the structure in the dimeric form. Furthermore, the binding of NADP+ induces conformational changes, leading to an open or closed conformation in the structure. Importantly, the binding of NADP+ does not disturb either the dimerization or oligomerization of the protein, a finding confirmed through both in vitro and in silico analysis. These findings shed light on the structural characteristics of WBm-ASDH and offer valuable insights for the development of new inhibitors specific to WBm, thereby contributing to the development of potential therapies for filarial parasitic infections.

Unveiling the intricacies of allosteric regulation in aspartate kinase from the Wolbachia endosymbiont of Brugia Malayi: Mechanistic and therapeutic insights.

Aspartate kinase (AK), an enzyme from the Wolbachia endosymbiont of Brugia malayi (WBm), plays a pivotal role in the bacterial cell wall and amino acid biosynthesis, rendering it an attractive candidate for therapeutic intervention. Allosteric inhibition of aspartate kinase is a prevalent mode of regulation across microorganisms and plants, often modulated by end products such as lysine, threonine, methionine, or meso-diaminopimelate. The intricate and diverse nature of microbial allosteric regulation underscores the need for rigorous investigation. This study employs a combined experimental and computational approach to decipher the allosteric regulation of WBmAK. Molecular Dynamics (MD) simulations elucidate that ATP (cofactor) and ASP (substrate) binding induce a closed conformation, promoting enzymatic activity. In contrast, the binding of lysine (allosteric inhibitor) leads to enzyme inactivation and an open conformation. The enzymatic assay demonstrates the optimal activity of WBmAK at 28 °C and a pH of 8.0. Notably, the allosteric inhibition study highlights lysine as a more potent inhibitor compared to threonine. Importantly, this investigation sheds light on the allosteric mechanism governing WBmAK and imparts novel insights into structure-based drug discovery, paving the way for the development of effective inhibitors against filarial pathogens.

Multilayer precision-based screening of potential inhibitors targeting Mycobacterium tuberculosis acetate kinase using in silico approaches.

Tuberculosis (TB), caused by Mycobacterium tuberculosis (MTB), remains a major threat to global health, with the emergence of multi-drug and extensively drug-resistant strains posing a serious challenge. Thereby, understanding the molecular basis of MTB virulence and disease pathogenesis is critical for developing effective therapeutic strategies. Targeting proteins involved in central metabolism has been recognized as a promising therapeutic approach to combat MTB. In this regard, the enzyme AckA of the acetate metabolic pathway which produces acetate from acetyl phosphate, is an important drug target for various pathogenic organisms. Therefore, this study aimed to identify potential AckA inhibitors through in silico methods, including molecular modeling, molecular dynamics simulation (MDS), and high-throughput virtual screening (HTVS) followed by ADMETox, MMGBSA, Density Functional Theory (DFT) calculations. HTVS of one million compounds from the ZINC database against AckA resulted in the top five hits (ZINC82048449, ZINC1219737510, ZINC1771921358, ZINC119699567, and ZINC1427100376) with better binding affinity and optimal binding free energy. MDS studies on complexes revealed that key residues, Asn195, Asp266, Phe267, Gly314, and Asn318 played a significant role in stable interactions of the top-ranked compounds to AckA. These outcomes provide insights into the optimal binding of the leads to inhibit the acetate pathway and aid in the rational design of novel therapeutic agents. Thus, the identified leads may act as promising compounds for targeting AckA and may serve as a potential therapeutic modality for treating TB. Our findings offer valuable insights into the inhibition of the acetate pathway, while also serving as a blueprint for rational drug design. The identified leads hold promise as compelling compounds for targeting AckA, thereby offering a potential therapeutic avenue for tackling TB. Thus, our study uncovers a pathway toward promising TB therapeutics by elucidating AckA inhibitors. By leveraging in silico methodologies, potent compounds that hold the potential to thwart AckA's role in MTB's acetate pathway have been unveiled. This breakthrough fosters optimism in the quest for novel and effective TB treatments, addressing a global health challenge with renewed vigor.

Exploring the antimicrobial potential of 4,5,7-trihydroxyflavanone (THF) against vancomycin-resistant Enterococcus gallinarum infections: in vitro and in silico investigations.

Enterococcus gallinarum and other Enterococcus species commonly inhabit the human gastrointestinal tract. While the pathogenicity of Enterococcus gallinarum remains incompletely understood, its infections are alarmingly severein humans, as evidenced by numerous cases. Formerly, Vancomycin was the preferred drug, but recent findings indicate that clinical isolates of Enterococcus gallinarum are resistant, leading to the emergence of vancomycin-resistant enterococci (VRE) strains. The escalation of drug resistance is often linked to overexpressed virulence factors, some of which are implicated in biofilm formation in Enterococcus infections. Henceforth, this research investigates the potential of phytocompounds to combat E. gallinarum infection, employing both in vitro and in silico methodologies. In vitro techniques were employed to assess the efficacy of various phytocompounds, ultimately identifying 4,5,7-trihydroxyflavanone (THF) as particularly effective in inhibiting microbial growth. THF displayed over 80% antibacterial activity at 200 µg/ml against E. gallinarum. Subsequent qualitative and quantitative hemolysin assays implicated hemolysin as a target of THF. Molecular docking analysis of THF and Hemolysin A revealed a strong binding affinity. Notably, residues Asn18, Asp85, and His199 formed hydrogen bonds, while His22 and His86 were involved in robust π-π stacking and π-cation interactions with THF. Overall, this study highlights THF's potential in combating E. gallinarum infections.Communicated by Ramaswamy H. Sarma.

Identification of potentially bioactive compounds from Blumea lacera essential oil by gas chromatography-mass spectroscopy and molecular docking studies for targeting inflammatory bowel disease.

Blumea lacera (Burm.f.) DC. (Asteraceae) is used in the traditional system of medicine for the treatment of inflammation or irritable bowel disease (IBD). In this study, B. lacera was collected from different geographical regions and oil was extracted by hydro-distillation and further chemo-profiled using GC-FID-MS. The major compounds identified were 2,5-dimethoxy-p-cymene (28.7-0.4%), ß-caryophyllene (25.5-0.5%), carvotanacetone (24.5-0.4%), chrysanthenone (21.9-9.8%) and 2,6-dimethyl phenol (11.4-1.8%). The constituents of B. lacera also showed marked qualitative and quantitative variations. The percent chemical similarity was observed to be in the range of 51.7% to 59.2% between the localities. Moreover, molecular modelling, membrane molecular dynamics simulations, target prediction were implemented to decipher the potential targets relevant to IBD. This inferred that all these major compounds could be potential drug moieties for treating IBD in terms of targeting h5HTR3A, thereby substantiating the traditional use of B. lacera for the treatment of IBD ailments.

Deciphering the structural and functional impact of missense mutations in Egr1-DNA interacting interface: an integrative computational approach.

Early growth response-1 (Egr1) is a zinc-finger transcription factor that plays a critical role in controlling cell growth, proliferation, differentiation, angiogenesis, and apoptosis. Egr1 is induced by many growth factors, cytokines, and stress signals and is also known to be involved in several pathological conditions like cancer, neurological and ocular disorders. The DNA binding domain of Egr1 is a highly conserved Cys2His2 (C2H2) zinc finger (ZNF) domain which specifically binds to GC-rich consensus sequence GcG (G/T) GGGCG and activates transcription. As the C2H2 domain specifically recognizes its DNA target, the mutations spanning this region shall perturb DNA recognition and may hinder transcription of target genes. Therefore, in this study, the missense mutations occurring specifically at the DNA binding domain (DBD) of Egr1 were probed by computational approaches involving in silico screening of pathogenic and functional mutants coupled with extensive molecular dynamics simulations, to determine the mutants that affect its structural stability and interactions with DNA. From the pathogenicity analysis of 38 missense mutations spanning Egr1-DBD, 17 were predicted as pathogenic, and 7 amongst these were found to have functional impact on Egr1. On combined analysis of molecular dynamics simulation, Residue interaction analysis and Egr1-DNA interaction analysis results, the mutants R371C and R375C showed least impact, whilst, H382R tend to increase the structural stability, whereas R360H, H390R, E393V, and H414Y conferred greater impact by altering the structural stability and DNA interactions. Hence, this study exposes the prospects of considering these 4 deleterious mutations for clinical significance, but needs further experimental validation.HighlightsEgr1's DNA binding domain is a highly conserved Cys2His2 (C2H2) zinc finger domain that specifically recognizes its DNA target.Mutations spanning in the DNA binding domain shall perturb DNA recognition and may hinder transcription.Among the missense mutations, mutants R360H, H390R, E393V, and H414Y were inferred to have a greater impact on Egr1 by altering the structural stability and DNA interactions.

Synthesis and Characterization of a Novel Peptide Targeting Human Tenon Fibroblast Cells To Modulate Fibrosis: An Integrated Empirical Approach.

Fibrosis is the primary factor influencing the prognosis of glaucoma post-trabeculectomy surgery, an eye condition characterized by increased intraocular pressure (IOP). Despite advancements in surgical procedures and aftercare, it continues to be a serious impediment. During the clinical intervention of scarring, fibrosis is managed by using topical application of combined antifibrotic drugs (mitomycin C). But still, scarring remains a key problem due to minimal drug penetration and nonbioavailability. In this study, we synthesized a cell-specific peptide for modulating scarring in human tenon fibroblasts undergoing epithelial-mesenchymal transition (EMT). The peptide was also conjugated with mitomycin C in order to investigate the effect of the drug conjugation on human tenon fibroblasts from the nanofiber composite system and to evaluate the fibrosis process. Peptide VRF2019 was identified using a subtractive proteomics approach, including solubility, cell penetration, and amphipathic properties. The peptide structure was determined using circular dichroism spectroscopy. The peptide and drug was conjugated using N-ethyl-N'-(3-(dimethylamino)propyl) carbodiimide/N-hydroxysuccinimide (EDC-NHS) chemistry, and the conjugation efficiency was evaluated using high-pressure liquid chromatography. The conjugated product and polycaprolactone (PCL) were electrospun to form a composite nanofiber, which was tested for cytotoxicity and drug release on human tenon fibroblast cells. The modeled VRF2019 peptide structure formed an α-helical structure with all residues spanning the allowed regions of the Ramachandran plot. Subsequent molecular dynamics simulations also demonstrated its membrane penetration potential. The peptide uptake was also studied in human tenon fibroblast cells. High-pressure liquid chromatography (HPLC) and mass spectrometry measurements confirmed peptide-drug conjugation and stability. Furthermore, scanning electron microscopy (SEM) investigation revealed the structure and size of the PCL composite nanofiber. We infer from early research that the PCL composite nanofiber matrix can greatly increase drug delivery and bioavailability.

Membrane dynamics simulation and virtual screening reveals potential dual natural inhibitors of endothelin receptors for targeting glaucomatous condition.

Glaucoma is the second leading cause of blindness in the world and is characterized by the loss of retinal ganglion cells (RGC) over a period of time, leading to complete blindness. Recently, endothelin has been identified as an important factor that influences intraocular pressure IOP, OBF, and direct RGC damage. Targeting the endothelin receptor signaling pathway in glaucoma is considered to be highly beneficial, as it can effectively modulate IOP, OBF, and RGC damage, the key factors which are essential to modulate the disease progression holistically. Currently, synthetic drugs like Bosentan, BQ-123, and prostaglandin analogues are available as endothelin receptor antagonists, which are extensively used in the treatment of cardiovascular and other conditions like systemic hypertension. However, the usage of these drugs in glaucoma is limited due to toxicity and poor bioavailability in the ocular milieu. Thus, there is a need for potential natural compounds as endothelin receptor antagonists that acts as dual inhibitors by targeting both ETA and ETB and are highly efficient with the least toxicity. Hence, this study is intended to prioritize endothelin receptor antagonists by structural bioinformatics approaches involving molecular modeling, molecular dynamics, and molecular docking studies. Subsequently, High throughput virtual screening (HTVS) vs. Natural compound databases targeting the optimal binding sites of both ETA and ETB. Following this, the common hits were subjected to binding free energy calculations (MMGBSA) and ADMETox analysis. Finally, the most potential hits were analyzed for MD based binding stability analysis and binding free energy. Similarly, the known synthetic inhibitors were also docked to the receptors and the results were analyzed. From this study, it was inferred that among the natural compounds dataset (8929 compounds), only 4 common compounds were identified as hits. Among these, only one compound ST075640 surpassed all the prioritization criteria including MMGBSA, ADMETox prediction, dual inhibitory potential (ETA & ETB), and also in structural comparative analysis with bosentan it showed similar efficiency. Thus, the validated hit shall prove to be effective in modulating endothelin mediated IOP, OBF, and RGC damage in glaucomatous condition.

Retinoschisis and Norrie disease: a missing link.

OBJECTIVE: Retinoschisis and Norrie disease are X-linked recessive retinal disorders caused by mutations in RS1 and NDP genes respectively. Both are likely to be monogenic and no locus heterogeneity has been reported. However, there are reports showing overlapping features of Norrie disease and retinoschisis in a NDP knock-out mouse model and also the involvement of both the genes in retinoschisis patients. Yet, the exact molecular relationships between the two disorders have still not been understood. The study investigated the association between retinoschisin (RS1) and norrin (NDP) using in vitro and in silico approaches. Specific protein-protein interaction between RS1 and NDP was analyzed in human retina by co-immunoprecipitation assay and MALDI-TOF mass spectrometry. STRING database was used to explore the functional relationship. RESULT: Co-immunoprecipitation demonstrated lack of a direct interaction between RS1 and NDP and was further substantiated by mass spectrometry. However, STRING revealed a potential indirect functional association between the two proteins. Progressively, our analyses indicate that FZD4 protein interactome via PLIN2 as well as the MAP kinase signaling pathway to be a likely link bridging the functional relationship between retinoschisis and Norrie disease.

Deciphering potential inhibitors targeting THI4 of Fusarium solani sp. to combat fungal keratitis: An integrative computational approach.

Mycotic keratitis is a fungal infection of corneal epithelium. It is more common in tropical and subtropical countries and is one of the leading causes of blindness. Many of the antifungal drugs have been less effective in treating this condition and certain drugs which are efficient and yet limited in use due to its extreme side effects. Hence, in this study an attempt is made to identify potential and least toxic antifungal inhibitors that targets thiamine thiazole synthase, a novel target for suppressing Fusarium solani subsp.pisi (Nectria haematococca MPVI) infections, to combat mycotic keratitis. Integrative computational approaches involving model refinement, molecular dynamics simulation and High throughput virtual screening (HTVS) were applied through integrative multi precision mode in order to identify potential inhibitors. Moreover, machine learning approach was also implemented to prioritize potential inhibitors that are ophthalmic adaptive, as well as antifungal molecule. From the NCI and Maybridge datasets, for HTVS only 209,872 compounds that surpassed ligand property filtration were considered, which resulted in 209 compounds after XP docking. Among the top 5 compounds from XP docking, on cumulative analysis only 2-(1-hydroxyethyl)-1,3-thiazole-4-carboxamide was prioritized as the most potential hit, as it showed higher order of significance in terms of binding affinity, structural stability and therapeutic relevance for the treatment of Mycotic keratitis. Thus, widening the scope for novel antifungal therapy in ophthalmic infections.

Structure-based drug target prioritisation and rational drug design for targeting Chlamydia trachomatis eye infections.

Chlamydia trachomatis (C.t) is a major causative of infectious blindness in world. It is a real challenge to combat Chlamydial infection as it is an intracellular pathogen. Hence, it is essential to determine the most potential targets of C.t in order to inhibit or suppress its virulence during its infectious phase. Thus, in this study, the highly expressed-cum-most essential genes reported through our earlier study were reprioritized by structure-based comparative binding site analysis with host proteome. Therefore, computational approaches involving molecular modelling, large-scale binding site prediction and comparison, molecular dynamics simulation studies were performed to narrow down the most potential targets. Furthermore, high-throughput virtual screening and ADMETox were also performed to identify potential hits that shall efficiently inhibit the prioritised targets. Hence, by this study we report Pyruvoyl-dependent arginine decarboxylase (PvlArgDC), DNA-repair protein (RecO) and porin (outer membrane protein) as the most viable targets of C.t which can be potentially targeted by compounds, NSC_13086, MFCD00276409, MFCD05662003, respectively. AbbreviationsC.tChlamydia trachomatisSTDSexually transmitted diseaseHTVSHigh-throughput virtual screeningADMEToxAbsorption, Distribution, Metabolism, Excretion and ToxicityPMPocketMatchMDMolecular Dynamics simulationSPStandard precisionXPExtra precisionMMGBSAMolecular mechanics energies combined with generalised Born and surface area continuum solvationOMPOuter membrane proteinPvlArgDCPyruvoyl-dependent arginine decarboxylaseRecORecombination protein O.Communicated by Ramaswamy H. Sarma.