Entamoeba histolytica Phosphoserine aminotransferase (EhPSAT): insights into the structure-function relationship

<p>Abstract</p> <p>Background</p> <p>Presence of phosphorylated Serine biosynthesis pathway upstream to the de novo cysteine biosynthesis pathway makes PSAT a crucial enzyme. Besides this, phoshoserine produced by the enzyme can also be taken up directly by cysteine synthase as a substrate. PSAT is a PLP dependent enzyme where the cofactor serves as an epicenter for functional catalysis with the active site architecture playing crucial role in optimum function of the enzyme.</p> <p>Findings</p> <p>EhPSAT is a homodimer of molecular mass 86 kDa. To understand the structural modulations associated with pH dependent changes in functional activity of EhPSAT detailed biophysical studies were carried out. pH alterations had no significant effect on the secondary structure, cofactor orientation and oligomeric configuration of the enzyme however, pH dependent compaction in molecular dimensions was observed. Most interestingly, a direct correlation between pH induced modulation of functional activity and orientation of Trp 101 present in the active site of the enzyme was observed. Sodium halides nullified the pH induced global changes in the enzyme, however differential effect of these salts on the active site microenvironment and functional activity of the enzyme was observed.</p> <p>Conclusions</p> <p>The study unequivocally demonstrates that pH induced selective modification of active site microenvironment and not global change in structure or oligomeric status of the enzyme is responsible for the pH dependent change in enzymatic activity of PSAT.</p

Can Mohalla Clinics be a Catalyst for Ayushman Bharat Pradhan Mantri Jan Arogya Yojna to achieve Universal Health Coverage in India?

BACKGROUND: Expenditure on health sector in India is one of the lowest public health expenditures in the world at just over 1% of GDP, with substantial disparities in population, infrastructure, availability and provision of services that define the Indian health care system. In a reform of Indian health systems approved by the Indian Government in March 2018, Ayushman Bharat Pradhan Mantri Jan Arogya Yojana (AB-PMJAY) was launched on 23rd September, 2018 to provide financial health insurance coverage of Rs. 5 lakhs per family per year for secondary and tertiary care hospitalizations to more than 10.74 million poor families that form the bottom 40% of population in India. OBJECTIVES: The objective of this paper is to discuss and analyze the benefits and weaknesses of the AB-PMJAY model and investigating the position that these clinics can play in (I) improving the provision of urban health facilities (II) resolving health inequities (III) and enhancing primary health care.MATERIALS AND METHOD: The authors explored different databases. Government portals, research publications on AB-PMJAY and Mohalla clinics (MC) to do in-depth analysis of various parameters.RESULTS: Although there are significant obstacles to the programme, by building impetus for program reform, AB-PMJAY offers an opportunity for the nation to resolve long-standing and ingrained shortcomings in governance, quality control and stewardship, and to accelerate India's development towards the stated UHC supply goal. The main achievement of these facilities in the last 18 months has been to introduce wellness to civic and political dialogue. CONCLUSION: Mohalla Clinics can prove as a major catalyst for Ayushman Bharat Pradhan Mantri Jan Arogya Yojna for health reforms and progress towards universal health coverage in India

Whole is Greater than Sum of Parts: Recognizing Scene Text Words

International audienceRecognizing text in images taken in the wild is a challenging problem that has received great attention in recent years. Previous methods addressed this problem by first detecting individual characters, and then forming them into words. Such approaches often suffer from weak character detections, due to large intra-class variations, even more so than characters from scanned documents. We take a different view of the problem and present a holistic word recognition framework. In this, we first represent the scene text image and synthetic images generated from lexicon words using gradient-based features. We then recognize the text in the image by matching the scene and synthetic image features with our novel weighted Dynamic Time Warping (wDTW) approach. We perform experimental analysis on challenging public datasets, such as Street View Text and ICDAR 2003. Our proposed method significantly outperforms our earlier work in Mishra et al. (CVPR 2012), as well as many other recent works, such as Novikova et al. (ECCV 2012), Wang et al. (ICPR 2012), Wang et al. (ICCV 2011)

Assembly of a dsRNA synthesizing complex: RNA-DEPENDENT RNA POLYMERASE 2 contacts the largest subunit of NUCLEAR RNA POLYMERASE IV

In plants, transcription of selfish genetic elements such as transposons and DNA viruses is suppressed by RNA-directed DNA methylation. This process is guided by 24-nt short-interfering RNAs (siRNAs) whose double-stranded precursors are synthesized by DNA-dependent NUCLEAR RNA POLYMERASE IV (Pol IV) and RNA-DEPENDENT RNA POLYMERASE 2 (RDR2). Pol IV and RDR2 coimmunoprecipitate, and their activities are tightly coupled, yet the basis for their association is unknown. Here, we show that an interval near the RDR2 active site contacts the Pol IV catalytic subunit, NRPD1, the largest of Pol IV's 12 subunits. Contacts between the catalytic regions of the two enzymes suggests that RDR2 is positioned to rapidly engage the free 3' ends of Pol IV transcripts and convert these single-stranded transcripts into double-stranded RNAs (dsRNAs)

Structural and Biochemical Features of Human Serum Albumin Essential for Eukaryotic Cell Culture

Serum albumin physically interacts with fatty acids, small molecules, metal ions, and several other proteins. Binding with a plethora of bioactive substances makes it a critical transport molecule. Albumin also scavenges the reactive oxygen species that are harmful to cell survival. These properties make albumin an excellent choice to promote cell growth and maintain a variety of eukaryotic cells under in vitro culture environment. Furthermore, purified recombinant human serum albumin is mostly free from impurities and modifications, providing a perfect choice as an additive in cell and tissue culture media while avoiding any regulatory constraints. This review discusses key features of human serum albumin implicated in cell growth and survival under in vitro conditions

An Explainable Model Using Graph-Wavelet for Predicting Biophysical Properties of Proteins and Measuring Mutational Effects

Proteins hold multispectral patterns of different kinds of physicochemical features of amino acids in their structures, which can help understand proteins&#x2019; behavior. Here, we propose a method based on the graph-wavelet transform of signals of features of amino acids in protein residue networks derived from their structures to achieve their abstract numerical representations. Such abstract representations of protein structures hand in hand with amino-acid features can be used for different purposes, such as modelling the biophysical property of proteins. Our method outperformed graph-Fourier and convolutional neural-network-based methods in predicting the biophysical properties of proteins. Even though our method does not predict deleterious mutations, it can summarize the effect of an amino acid based on its location and neighbourhood in protein-structure using graph-wavelet to estimate its influence on the biophysical property of proteins. Such an estimate of the influence of amino-acid has the potential to explain the mechanism of the effect of deleterious non-synonymous mutations. Thus, our approach can reveal patterns of distribution of amino-acid properties in the structure of the protein in the context of a biophysical property for better classification and more insightful understanding

Patterns of transcription factor binding and epigenome at promoters allow interpretable predictability of multiple functions of non-coding and coding genes

Understanding the biological roles of all genes only through experimental methods is challenging. A computational approach with reliable interpretability is needed to infer the function of genes, particularly for non-coding RNAs. We have analyzed genomic features that are present across both coding and non-coding genes like transcription factor (TF) and cofactor ChIP-seq (823), histone modifications ChIP-seq (n = 621), cap analysis gene expression (CAGE) tags (n = 255), and DNase hypersensitivity profiles (n = 255) to predict ontology-based functions of genes. Our approach for gene function prediction was reliable (>90% balanced accuracy) for 486 gene-sets. PubMed abstract mining and CRISPR screens supported the inferred association of genes with biological functions, for which our method had high accuracy. Further analysis revealed that TF-binding patterns at promoters have high predictive strength for multiple functions. TF-binding patterns at the promoter add an unexplored dimension of explainable regulatory aspects of genes and their functions. Therefore, we performed a comprehensive analysis for the functional-specificity of TF-binding patterns at promoters and used them for clustering functions to reveal many latent groups of gene-sets involved in common major cellular processes. We also showed how our approach could be used to infer the functions of non-coding genes using the CRISPR screens of coding genes, which were validated using a long non-coding RNA CRISPR screen. Thus our results demonstrated the generality of our approach by using gene-sets from CRISPR screens. Overall, our approach opens an avenue for predicting the involvement of non-coding genes in various functions

Matching queried single-cell open-chromatin profiles to large pools of single-cell transcriptomes and epigenomes for reference supported analysis

The true benefits of large single-cell transcriptome and epigenome data sets can be realized only with the development of new approaches and search tools for annotating individual cells. Matching a single-cell epigenome profile to a large pool of reference cells remains a major challenge. Here, we present scEpiSearch, which enables searching, comparison, and independent classification of single-cell open-chromatin profiles against a large reference of single-cell expression and open-chromatin data sets. Across performance benchmarks, scEpiSearch outperformed multiple methods in accuracy of search and low-dimensional coembedding of single-cell profiles, irrespective of platforms and species. Here we also demonstrate the unconventional utilities of scEpiSearch by applying it on single-cell epigenome profiles of K562 cells and samples from patients with acute leukaemia to reveal different aspects of their heterogeneity, multipotent behavior, and dedifferentiated states. Applying scEpiSearch on our single-cell open-chromatin profiles from embryonic stem cells (ESCs), we identified ESC subpopulations with more activity and poising for endoplasmic reticulum stress and unfolded protein response. Thus, scEpiSearch solves the nontrivial problem of amalgamating information from a large pool of single cells to identify and study the regulatory states of cells using their single-cell epigenomes

Structure and RNA template requirements of Arabidopsis RNA-DEPENDENT RNA POLYMERASE 2

RNA-dependent RNA polymerases play essential roles in RNA-mediated gene silencing in eukaryotes. In Arabidopsis, RNA-DEPENDENT RNA POLYMERASE 2 (RDR2) physically interacts with DNA-dependent NUCLEAR RNA POLYMERASE IV (Pol IV) and their activities are tightly coupled, with Pol IV transcriptional arrest, induced by the nontemplate DNA strand, somehow enabling RDR2 to engage Pol IV transcripts and generate double-stranded RNAs. The double-stranded RNAs are then released from the Pol IV-RDR2 complex and diced into short-interfering RNAs that guide RNA-directed DNA methylation and silencing. Here we report the structure of full-length RDR2, at an overall resolution of 3.1 Å, determined by cryoelectron microscopy. The N-terminal region contains an RNA-recognition motif adjacent to a positively charged channel that leads to a catalytic center with striking structural homology to the catalytic centers of multisubunit DNA-dependent RNA polymerases. We show that RDR2 initiates 1 to 2 nt internal to the 3' ends of its templates and can transcribe the RNA of an RNA/DNA hybrid, provided that 9 or more nucleotides are unpaired at the RNA's 3' end. Using a nucleic acid configuration that mimics the arrangement of RNA and DNA strands upon Pol IV transcriptional arrest, we show that displacement of the RNA 3' end occurs as the DNA template and nontemplate strands reanneal, enabling RDR2 transcription. These results suggest a model in which Pol IV arrest and backtracking displaces the RNA 3' end as the DNA strands reanneal, allowing RDR2 to engage the RNA and synthesize the complementary strand

From PROTAC to inhibitor: Structure-guided discovery of potent and orally bioavailable BET inhibitors

An X-ray structure of a CLICK chemistry-based BET PROTAC bound to BRD2(BD2) inspired synthesis of JQ1 derived heterocyclic amides. This effort led to the discovery of potent BET inhibitors displaying overall improved profiles when compared to JQ1 and birabresib. A thiadiazole derived 1q (SJ1461) displayed excellent BRD4 and BRD2 affinity and high potency in the panel of acute leukaemia and medulloblastoma cell lines. A structure of 1q co-crystalised with BRD4-BD1 revealed polar interactions with the AZ/BC loops, in particular with Asn140 and Tyr139, rationalising the observed affinity improvements. In addition, exploration of pharmacokinetic properties of this class of compounds suggest that the heterocyclic amide moiety improves drug-like features. Our study led to the discovery of potent and orally bioavailable BET inhibitor 1q (SJ1461) as a promising candidate for further development