Inhibiting the Keap1/Nrf2 Protein-Protein Interaction with Protein-Like Polymers.

Successful and selective inhibition of the cytosolic protein-protein interaction (PPI) between nuclear factor erythroid 2-related factor 2 (Nrf2) and Kelch-like ECH-associating protein 1 (Keap1) can enhance the antioxidant response, with the potential for a therapeutic effect in a range of settings including in neurodegenerative disease (ND). Small molecule inhibitors have been developed, yet many have off-target effects, or are otherwise limited by poor cellular permeability. Peptide-based strategies have also been attempted to enhance specificity, yet face challenges due to susceptibility to degradation and lack of cellular penetration. Herein, these barriers are overcome utilizing a polymer-based proteomimetics. The protein-like polymer (PLP) consists of a synthetic, lipophilic polymer backbone displaying water soluble Keap1-binding peptides on each monomer unit forming a brush polymer architecture. The PLPs are capable of engaging Keap1 and displacing the cellular protective transcription factor Nrf2, which then translocates to the nucleus, activating the antioxidant response element (ARE). PLPs exhibit increased Keap1 binding affinity by several orders of magnitude compared to free peptides, maintain serum stability, are cell-penetrant, and selectively activate the ARE pathway in cells, including in primary cortical neuronal cultures. Keap1/Nrf2-inhibitory PLPs have the potential to impact the treatment of disease states associated with dysregulation of oxidative stress, such as NDs.

Zirconia Facts and Perspectives for Biomaterials in Dental Implantology.

Dental implantology has witnessed remarkable advancements in recent years, and zirconia has emerged as a prominent biomaterial for dental implant applications. This review explores the multifaceted aspects of zirconia, focusing on its properties, processing methods, biocompatibility, mechanical performance, and clinical applications. Over the past few decades, the most popular choice of material for dental implantology has been titanium which has been found to have the highest success rate of implant treatment. However, recently, it has been observed that zirconia might replace titanium and eventually emerge as one of the gold-standard materials of dental implants. Analysis of biomechanical sciences and biomaterial sciences provides an opportunity for the refinement of design and material notions for surgical implants. However, the most important aspect and prime concern is how tissue at the implant site responds to biomechanical disturbances caused by foreign materials. The literature revealed that zirconia has certain characteristics that make it an excellent material for implants, including biocompatibility and osseointegration which depicts positive soft tissue response with low plaque affinity as well as aesthetics owing to light transmission and color. Additionally, this review discusses the current challenges and prospects of zirconia in dental implantology as well as aims to provide dental professionals and researchers with a comprehensive understanding of zirconia's potential as a biomaterial in dental implantology. The present overview of available literature intends to highlight and explore the biological characteristics of zirconia for applications in dental implantology. However, research is urgently required to fill in gaps over time for clinical assessments of all zirconia implants, consequently, the implementation of hybrid systems (a titanium screw with a zirconia collar) has recently been suggested.

The Prevalence of Dental Fear and Its Relationship to Dental Caries and Gingival Diseases Among School Children in Wardha.

Background Children who are afraid of the dentist have terrible behavioral effects, and one of those effects is that they have a preconceived concept that getting treatment would be unpleasant. Such fear and anxiety can lead to avoidance of dental care. These patients must be recognized and their concerns addressed as soon as possible. It is, therefore, important to highlight the connection between the constructs that target the development of dental fear and anxiety, including its outcome in children concerning the prevalence of dental diseases. Aims and objectives To assess the prevalence of dental anxiety and its correlation to dental caries and gingivitis in students in Wardha aged six to 12 years old. Methods Two hundred schoolchildren between the ages of six and 12 were chosen at random, with 100 boys and 100 girls. Children had an oral examination utilizing the decayed, missing, and filled teeth (DMFT) Index as well as the Loe and Silness gingival index (GI), as well as a modified version of the dental fear survey questionnaire. Results In the study population, the prevalence of low to moderate "general dental fear" was 47%, whereas the frequency of high dental fear was 14%. The mean DMFT (1.80 ± 1.76) and GI (1.04 ± 0.52) of boys did not differ substantially from the DMFT (1.94 ± 81.02) and GI (0.97 ± 0.53) of girls (P > 0.05). Conclusion In terms of DMFT and gingival scores, there was not any obvious distinction between male and female children. Additionally, there was no connection found between "general dental fear" and either the DMFT or GI scores. Dental fear scores decreased with increasing age.

The Ndc80-Cdt1-Ska1 complex is a central processive kinetochore-microtubule coupling unit.

It is known that microtubule-binding proteins including the Ska1 complex and the DNA replication licensing factor, Cdt1, enable the kinetochore-localized Ndc80 complex to form robust kinetochore-microtubule attachments. However, it is not clear how the Ndc80 complex is stably coupled to dynamic spindle microtubule plus-ends. Here, we have developed a conditional auxin-inducible degron approach to reveal a function for Cdt1 in chromosome segregation and kinetochore-microtubule interactions that is separable from its role in DNA replication licensing. Further, we demonstrate that a direct interaction between Cdt1 and Ska1 is required for recruiting Cdt1 to kinetochores and spindle microtubules. Cdt1 phosphorylation by Cdk1 kinase is critical for Ska1 binding, kinetochore-microtubule attachments, and mitotic progression. Furthermore, we show that Cdt1 synergizes with Ndc80 and Ska1 for microtubule binding, including forming a diffusive, tripartite Ndc80-Cdt1-Ska1 complex that can processively track dynamic microtubule plus-ends in vitro. Taken together, our data identify the Ndc80-Cdt1-Ska1 complex as a central molecular unit that can promote processive bidirectional tip-tracking of microtubules by kinetochores.

Integrative big transcriptomics data analysis implicates crucial role of MUC13 in pancreatic cancer.

Big data analysis holds a considerable influence on several aspects of biomedical health science. It permits healthcare providers to gain insights from large and complex datasets, leading to improvements in the understanding, diagnosis, medication, and restraint of pathological conditions including cancer. The incidences of pancreatic cancer (PanCa) are sharply rising, and it will become the second leading cause of cancer related deaths by 2030. Various traditional biomarkers are currently in use but are not optimal in sensitivity and specificity. Herein, we determine the role of a new transmembrane glycoprotein, MUC13, as a potential biomarker of pancreatic ductal adenocarcinoma (PDAC) by using integrative big data mining and transcriptomic approaches. This study is helpful to identify and appropriately segment the data related to MUC13, which are scattered in various data sets. The assembling of the meaningful data, representation strategy was used to investigate the MUC13 associated information for the better understanding regarding its structural, expression profiling, genomic variants, phosphorylation motifs, and functional enrichment pathways. For further in-depth investigation, we have adopted several popular transcriptomic methods like DEGseq2, coding and non-coding transcript, single cell seq analysis, and functional enrichment analysis. All these analyzes suggest the presence of three nonsense MUC13 genomic transcripts, two protein transcripts, short MUC13 (s-MUC13, non-tumorigenic or ntMUC13), and long MUC13 (L-MUC13, tumorigenic or tMUC13), several important phosphorylation sites in tMUC13. Altogether, this data confirms that importance of tMUC13 as a potential biomarker, therapeutic target of PanCa, and its significance in pancreatic pathobiology.

Multiple Protein Biomarkers and Different Treatment Strategies for Colorectal Carcinoma: A Comprehensive Prospective.

In this review, we emphasized important biomarkers, pathogenesis, and newly developed therapeutic approaches in the treatment of colorectal cancer (CRC). This includes a complete description of small-molecule inhibitors, phytopharmaceuticals with antiproliferative potential, monoclonal antibodies for targeted therapy, vaccinations as immunotherapeutic agents, and many innovative strategies to intervene in the interaction of oncogenic proteins. Many factors combine to determine the clinical behavior of colorectal cancer and it is still difficult to comprehend the molecular causes of a person's vulnerability to CRC. It is also challenging to identify the causes of the tumor's onset, progression, and responsiveness or resistance to antitumor treatment. Current recommendations for targeted medications are being updated by guidelines throughout the world in light of the growing number of high-quality clinical studies. So, being concerned about the aforementioned aspects, we have tried to present a summarized pathogenic view, including a brief description of biomarkers and an update of compounds with their underlying mechanisms that are currently under various stages of clinical testing. This will help to identify gaps or shortfalls that can be addressed in upcoming colorectal cancer research.

A Study to Evaluate the Effect of Opportunistic Salpingectomy on Ovarian Reserve and Function.

Background: There are conflicting reports on status of ovarian function after hysterectomy and opportunistic salpingectomy in premenopausal women. The present study was undertaken to understand the effect of salpingectomy done at the time of hysterectomy on ovarian reserve and function as measured by serum AMH and FSH levels before and after the surgery. Methods: This was a prospective study conducted on 60 women who underwent hysterectomy at our tertiary care centre, Shri Guru Ram Rai Institute of medical and health sciences, Dehradun, from January 2020 to September 2021. Serum AMH and FSH levels were monitored preoperatively and 3 months postoperatively in patients undergoing hysterectomy with bilateral salpingectomy and hysterectomy without salpingectomy. Results: The mean age of the patients was 41.83 yrs in group 1 and 43.73 yrs in group 2 [p value = 0.078]. Most common indication of hysterectomy was AUB-L in both the groups (86% and 80%, respectively). Mean operative time was 115.50 min in group 1 and 114.40 min in group 2 [p value = 0.823]. Mean intra-operative blood loss was 214 ml in group 1 and 199.33 ml in group 2 [p value = 0.087]. Serum AMH and FSH were insignificantly decreased in both the groups post-operatively after 3 months, and the difference between both groups was also not statistically significant. Conclusion: Salpingectomy done at the time of hysterectomy for benign indications with preservation of ovaries did not have any short-term adverse effects on ovarian reserve and function.

Structural Insight on GPR119 Agonist as Potential Therapy for Type II Diabetes: A Comprehensive Review.

Diabetes Mellitus (DM) is a long-term metabolic condition that is characterized by excessive blood glucose. DM is the third most death-causing disease, leading to retinopathy, nephropathy, loss of vision, stroke, and cardiac arrest. Around 90% of the total cases of diabetic patients have Type II Diabetes Mellitus (T2DM). Among various approaches for the treatment of T2DM. G proteincoupled receptors (GPCRs) 119 have been identified as a new pharmacological target. GPR119 is distributed preferentially in the pancreas ß-cells and gastrointestinal tract (enteroendocrine cells) in humans. GPR119 receptor activation elevates the release of incretin hormones such as Glucagon-Like Peptide (GLP1) and Glucose Dependent Insulinotropic Polypeptide (GIP) from intestinal K and L cells. GPR119 receptor agonists stimulate intracellular cAMP production via Gαs coupling to adenylate cyclase. GPR119 has been linked to the control of insulin release by pancreatic ß-cells, as well as the generation of GLP-1 by enteroendocrine cells in the gut, as per in vitro assays. The dual role of the GPR119 receptor agonist in the treatment of T2DM leads to the development of a novel prospective anti-diabetic drug and is thought to have decreased the probability of inducing hypoglycemia. GPR119 receptor agonists exert their effects in one of two ways: either by promoting glucose absorption by ß-cells, or by inhibiting α-cells' ability to produce glucose. In this review, we summarized potential targets for the treatment of T2DM with special reference to GPR119 along with its pharmacological effects, several endogenous as well as exogenous agonists, and its pyrimidine nucleus containing synthetic ligands.

In-Depth Comparison of Genetic Variants Demonstrates a Close Relationship Between Invasive and Intraductal Components of Prostate Cancer.

Intraductal carcinoma (IDC) of the prostate is often associated with concurrent high-grade invasive prostate cancer (PCa) and poor clinical outcomes. In this context, IDC is thought to represent the retrograde spread of invasive prostatic adenocarcinoma into the acini and ducts. Prior studies have demonstrated a concordance of PTEN loss and genomic instability between the IDC and high-grade invasive components of PCa, but larger genomic association studies to solidify our understanding of the relationship between these 2 lesions are lacking. Here, we evaluate the genomic relationship between duct-confined (high-grade prostatic intraepithelial neoplasia and IDC) and invasive components of high-grade PCa using genetic variants generated by whole exome sequencing. High-grade prostatic intraepithelial neoplasia and IDC were laser-microdissected, and PCa and nonneoplastic tissue was manually dissected from 12 radical prostatectomies. A targeted next-generation sequencing panel was used to identify disease-relevant variants. Additionally, the degree of overlap between adjacent lesions was determined by comparing exome-wide variants detected using whole exome sequencing data. Our results demonstrate that IDC and invasive high-grade PCa components show common genetic variants and copy number alterations. Hierarchical clustering of genome-wide variants suggests that in these tumors, IDC is more closely related to the high-grade invasive components of the tumor compared with high-grade prostatic intraepithelial neoplasia. In conclusion, this study reinforces the concept that, in the context of high-grade PCa, IDC likely represents a late event associated with tumor progression.

Structural Perspective of Benzophenones Targeting Tubulin as Anticancer Agents.

Cancer is the leading cause of death and the most significant determinant of life expectancy in almost every country in this twenty-first century. According to the World Health Organization (WHO), cancer is responsible for the leading cause of death globally. Benzophenone derivatives are found in a variety of naturally occurring compounds which are known to be pharmacologically efficacious against a variety of diseases, including cancer. Microtubules are thought to be a good target for cancer chemotherapies. Microtubule polymerization and depolymerization are induced by a variety of natural, synthetic, and semisynthetic chemicals having a benzophenone nucleus, affecting tubulin dynamics. Several medications that affect microtubule dynamics are in various stages of clinical trials, including Combretastatins (phase II), Vincristine (clinically approved), Paclitaxel (in clinical usage), and epothilone (phase III), and only a few have been patented. Benzophenone derivatives target the colchicine binding site of microtubules, damage them and cause cell cycle arrest in the G2-M phase. Belonging to this class of molecules, phenstatin, a potent inhibitor of tubulin polymerization, has shown strongly inhibit cancer cell growth and arrest the G2/M phase of the cell cycle by targeting the colchicine binding site of microtubules. In the present manuscript, we described the benzophenone as tubulin polymerization inhibitors, their Structure-Activity Relationships (SARs) and molecular docking studies that reveal its binding affinity with the colchicine binding site.

Multiple strategies for the treatment of invasive breast carcinoma: A comprehensive prospective.

In this review, we emphasize on evolving therapeutic strategies and advances in the treatment of breast cancer (BC). This includes small-molecule inhibitors under preclinical and clinical investigation, phytoconstituents with antiproliferative potential, targeted therapies as antibodies and antibody-drug conjugates (ADCs), vaccines as immunotherapeutic agents and peptides as a novel approach inhibiting the interaction of oncogenic proteins. We provide an update of molecules under different phases of clinical investigation which aid in the identification of loopholes or shortcomings that can be overcomed with future breast cancer research.

Deiodinase-3 is a thyrostat to regulate podocyte homeostasis.

BACKGROUND: Nephrotic syndrome (NS) is associated with kidney podocyte injury and may occur as part of thyroid autoimmunity such as Graves' disease. Therefore, the present study was designed to ascertain if and how podocytes respond to and regulate the input of biologically active thyroid hormone (TH), 3,5,3'-triiodothyronine (T3); and also to decipher the pathophysiological role of type 3 deiodinase (D3), a membrane-bound selenoenzyme that inactivates TH, in kidney disease. METHODS: To study D3 function in healthy and injured (PAN, puromycin aminonucleoside and LPS, Lipopolysaccharide-mediated) podocytes, immunofluorescence, qPCR and podocyte-specific D3 knockout mouse were used. Surface plasmon resonance (SPR), co-immunoprecipitation and Proximity Ligation Assay (PLA) were used for the interaction studies. FINDINGS: Healthy podocytes expressed D3 as the predominant deiodinase isoform. Upon podocyte injury, levels of Dio3 transcript and D3 protein were dramatically reduced both in vitro and in the LPS mouse model of podocyte damage. D3 was no longer directed to the cell membrane, it accumulated in the Golgi and nucleus instead. Further, depleting D3 from the mouse podocytes resulted in foot process effacement and proteinuria. Treatment of mouse podocytes with T3 phenocopied the absence of D3 and elicited activation of αvß3 integrin signaling, which led to podocyte injury. We also confirmed presence of an active thyroid stimulating hormone receptor (TSH-R) on mouse podocytes, engagement and activation of which resulted in podocyte injury. INTERPRETATION: The study provided a mechanistic insight into how D3-αvß3 integrin interaction can minimize T3-dependent integrin activation, illustrating how D3 could act as a renoprotective thyrostat in podocytes. Further, injury caused by binding of TSH-R with TSH-R antibody, as found in patients with Graves' disease, explained a plausible link between thyroid disorder and NS. FUNDING: This work was supported by American Thyroid Association (ATA-2018-050.R1).

Renal cell markers: lighthouses for managing renal diseases.

Kidneys, one of the vital organs in our body, are responsible for maintaining whole body homeostasis. The complexity of renal function (e.g., filtration, reabsorption, fluid and electrolyte regulation, and urine production) demands diversity not only at the level of cell types but also in their overall distribution and structural framework within the kidney. To gain an in depth molecular-level understanding of the renal system, it is imperative to discern the components of kidney and the types of cells residing in each of the subregions. Recent developments in labeling, tracing, and imaging techniques have enabled us to mark, monitor, and identify these cells in vivo with high efficiency in a minimally invasive manner. In this review, we summarize different cell types, specific markers that are uniquely associated with those cell types, and their distribution in the kidney, which altogether make kidneys so special and different. Cellular sorting based on the presence of certain proteins on the cell surface allowed for the assignment of multiple markers for each cell type. However, different studies using different techniques have found contradictions in cell type-specific markers. Thus, the term "cell marker" might be imprecise and suboptimal, leading to uncertainty when interpreting the data. Therefore, we strongly believe that there is an unmet need to define the best cell markers for a cell type. Although the compendium of renal-selective marker proteins presented in this review is a resource that may be useful to researchers, we acknowledge that the list may not be necessarily exhaustive.

From Infancy to Fancy: A Glimpse into the Evolutionary Journey of Podocytes in Culture.

Podocytes are critical components of the filtration barrier and responsible for maintaining healthy kidney function. An assault on podocytes is generally associated with progression of chronic glomerular diseases. Therefore, podocyte pathophysiology is a favorite research subject for nephrologists. Despite this, podocyte research has lagged because of the unavailability of techniques for culturing such specialized cells ex vivo in quantities that are adequate for mechanistic studies. In recent years, this problem was circumvented by the efforts of researchers, who successfully developed several in vitro podocyte cell culture model systems that paved the way for incredible discoveries in the field of nephrology. This review sets us on a journey that provides a comprehensive insight into the groundbreaking breakthroughs and novel technologic advances made in the field of podocyte cell culture so far, beginning from its inception, evolution, and progression. In this study, we also describe in detail the pros and cons of different models that are being used to culture podocytes. Our extensive and exhaustive deliberation on the status of podocyte cell culture will facilitate researchers to choose wisely an appropriate model for their own research to avoid potential pitfalls in the future.

Potential target identification for breast cancer and screening of small molecule inhibitors: A bioinformatics approach.

In the current study, we investigated the role of PAK1 (P21 (RAC1) Activated Kinase 1) gene in breast cancer and to this end, we performed differential gene expression analysis of PAK1 in breast cancer tissues compared to the normal adjacent tissue. We also studied its significance in protein-protein interaction (PPI) network, and analysed biological pathways, cellular processes, and role of PAK1 in different diseases. We found PAK1 to have significant role in breast cancer pathways such as integrin signaling, axonal guidance signaling, signaling by Rho family GTPases, ERK5 signaling. Additionally, it has been found as hub gene in PPI network, suggesting its possible regulatory role in breast carcinogenesis. Moreover, PAK1 had role in progression of various diseases as neoplasia, tumorigenesis, lymphatic neoplasia. Thereby, PAK1 can be used as a therapeutic target in breast cancer. Further, we put our efforts in identification of potential small molecules inhibitors against PAK1 by developing a composite virtual screening protocol involving molecular dynamics (MD) and molecular docking. The chemical library of compounds from NCI diversity sets, Pubchem and eMolecules were screened against PAK1 protein and hits which showed good binding affinity were considered for MD simulation study. Moreover, to assess binding of selected hits, MMGBSA (Molecular Mechanics-Generalized Born Surface Area) analysis was performed using AMBER (Assisted Model Building with Energy Refinement) package. MMGBSA calculations exhibited that the identified ligands showed good binding affinity with PAK1. HighlightsThe PAK1 has been found to be upregulated in breast cancer samples and is a potential oncogene playing role in different cellular functions and processes.The molecular docking studies revealed ligands showed good binding affinity towards PAK1 protein.The residues Glu345, Leu347, Thr406, Asp299, Asp393 and Gly350 were found to make H-bond interactions with small molecule inhibitors.The residues Ile276, Val284, Ala297, Tyr346, Leu396 and Asp407 were found to make hydrophobic interactions.The RMSD analysis confirmed stability of complexes throughout 40 ns production period.The MD simulations studies revealed the binding site flexibility, binding free energy of complexes and per-residue contribution in ligand binding.Communicated by Ramaswamy H. Sarma.

The Recent Development of Piperazine and Piperidine Derivatives as Antipsychotic Agents.

Schizophrenia is a chronic neuropsychiatric disorder that affects nearly 1% of the global population. There are various anti-psychotic drugs available for the treatment of schizophrenia, but they have certain side effects; therefore, there is a need to explore and develop novel potential lead compounds against schizophrenia. The currently available drugs e.g. typical and atypical antipsychotics act on different dopamine and serotonin receptors and as per literature reports, various piperidine and piperazine derivatives have shown promising activity against these receptors. When different heterocyclic groups are attached to basic piperidine and piperazine rings, the antipsychotic activity is greatly potentiated. In this direction, various antipsychotic drugs have been synthesized at the laboratory level, and few are under clinical trial studies, such as Lu AE58054, PF-04802540, ORG25935, DMXB-A, Bitopertin, and ABT-126. In the present review, we include the studies related to the effect of different substituents on piperidine/piperazine derivatives and their anti-psychotic activity. Various series of synthesized compounds by other researchers with piperidine/piperazine nucleus have been reviewed and diagrammatically represented in the form of SAR (structure-activity relationships), which will help the scientists for the development of potential lead compounds.

Current Insights into the Chemistry and Antitubercular Potential of Benzimidazole and Imidazole Derivatives.

Tuberculosis is a disease caused by Mycobacterium tuberculosis (Mtb), affecting millions of people worldwide. The emergence of drug resistance is a major problem in the successful treatment of tuberculosis. Due to the commencement of MDR-TB (multi-drug resistance) and XDR-TB (extensively drug resistance), there is a crucial need for the development of novel anti-tubercular agents with improved characteristics such as low toxicity, enhanced inhibitory activity and short duration of treatment. In this direction, various heterocyclic compounds have been synthesized and screened against Mycobacterium tuberculosis. Among them, benzimidazole and imidazole containing derivatives have been found to have potential anti-tubercular activity. The present review focuses on various imidazole and benzimidazole derivatives (from 2015-2019) with their structure-activity relationships in the treatment of tuberculosis.

An exhaustive perspective on structural insights of SGLT2 inhibitors: A novel class of antidiabetic agent.

Diabetes mellitus is the global health issue and become an alarming threat in the modern era where human lifestyle gets compromised with modernization. According to the latest statistical report 2020, USA has 9.47% (31 million among 32.72 cr), China has 8.3% (116.4 million among 139.27 cr) and India has 5.6% (77 million among 135.26 cr) of the diabetic people, indicating that diabetes is more prevailing in developed countries as compared to the developing countries. The number of diabetic patients is rising day by day at a tremendous rate and soon it may affect each and every person in a family. So, there is an urgent need to develop novel entities that can meet the scarcity of present antidiabetic agents. In the last few decades, the sodium-glucose co-transporter 2 (SGLT2) has emerged as a prominent target for the treatment of Type 2 diabetes mellitus due to its novel mechanism of action & no involvement in insulin signaling pathway. Most of the inhibitors that target SGLT2 contain three basic moieties: glucose, two benzene rings (one is connected with glucose and the other with methylene), and the methylene bridge which are similar to dapagliflozin. Several SGLT2 inhibitors and their derivatives such as remogliflozin etabonate (phase-II), sotagliflozin (phase-III) and bexagliflozin (phase-III) are under different phases of clinical trial studies and some have been patented. The present review is focused on SGLT2 inhibitors, structure activity relationships (SARs) of dapagliflozin and its several analogues for their binding affinity with SGLT2. We have also presented and summarized the efforts made by various researchers in terms of the synthesis of various dapagliflozin derivatives till date.

Ligand and structure based virtual screening of chemical databases to explore potent small molecule inhibitors against breast invasive carcinoma using recent computational technologies.

Breast carcinoma is the most common invasive cancer to affect the women in the North America and the world. Cancer of breast is the number one cancer overall with estimated 1.5 lakh new cases during 2016. The success of the current endocrine therapies is often limited due to the development of resistance. Therefore, there is a need to develop new lead compounds for breast cancer treatment. As 70% of breast carcinoma is ER+, and it is well known previously that estrogen receptor alpha (ERα) is overexpressed in ER + cases, so in the current work we attempt to develop some novel potent analogues against ERα. To achieve this, we have adopted an integrative computational approach that involves multiple sequence alignment, virtual screening (ligand and structure based), molecular docking, fingerprint based clustering and molecular dynamics simulation. The approach envisaged vital information about the binding site residues, conserved sequence among different species, ligand and protein conformations, binding energy of compound to bind into the active site of the receptor. Molecular docking analysis revealed that some analogues exhibited significant binding towards ERα. The top docked complexes showing good docking scores, hydrogen bond and hydrophobic interactions were selected for molecular dynamics simulation studies. RMSD revealed that the systems were quite stable with RMSD value below 3 Å. The RMSF analysis calculated residue wise fluctuations and revealed that the residues are flexible enough to interact with the ligand. The residue at C-terminal showed more flexibility as compared to other residues. To confirm binding of these analogues, MMGBSA analysis was performed which revealed binding energy of the ligands. Further, per-residue decomposition energy analysis revealed that Glu353, Leu346, Leu387 and Arg394 contributed towards ligand binding. The results visibly indicated that MMGBSA can act as filter in virtual screening experiments and play a major role in facilitating drug discovery.

N-terminal autoprocessing and acetylation of multifunctional-autoprocessing repeats-in-toxins (MARTX) Makes Caterpillars Floppy-like effector is stimulated by adenosine diphosphate (ADP)-Ribosylation Factor 1 in advance of Golgi fragmentation.

Studies have successfully elucidated the mechanism of action of several effector domains that comprise the multifunctional-autoprocessing repeats-in-toxins (MARTX) toxins of Vibrio vulnificus. However, the biochemical linkage between the cysteine proteolytic activity of Makes Caterpillars Floppy (MCF)-like effector and its cellular effects remains unknown. In this study, we identify the host cell factors that activate in vivo and in vitro MCF autoprocessing as adenosine diphosphate (ADP)-Ribosylation Factor 1 (ARF1) and ADP-Ribosylation Factor 3 (ARF3). Autoprocessing activity is enhanced when ARF1 is in its active [guanosine triphosphate (GTP)-bound] form compared to the inactive [guanosine diphosphate (GDP)-bound] form. Subsequent to auto-cleavage, MCF is acetylated on its exposed N-terminal glycine residue. Acetylation apparently does not dictate subcellular localization as MCF is found localized throughout the cell. However, the cleaved form of MCF gains the ability to bind to the specialized lipid phosphatidylinositol 5-phosphate enriched in Golgi and other membranes necessary for endocytic trafficking, suggesting that a fraction of MCF may be subcellularly localized. Traditional thin-section electron microscopy, high-resolution cryoAPEX localization, and fluorescent microscopy show that MCF causes Golgi dispersal resulting in extensive vesiculation. In addition, host mitochondria are disrupted and fragmented. Mass spectrometry analysis found no reproducible modifications of ARF1 suggesting that ARF1 is not post-translationally modified by MCF. Further, catalytically active MCF does not stably associate with ARF1. Our data indicate not only that ARF1 is a cross-kingdom activator of MCF, but also that MCF may mediate cytotoxicity by directly targeting another yet to be identified protein. This study begins to elucidate the biochemical activity of this important domain and gives insight into how it may promote disease progression.