Factors determining institutional delivery in eastern part of India.

Objective: Institutional delivery (ID) plays a vital role for determining the outcome of pregnancy and ensuring care to mother and newborn. The present study aims to find out the correlates of ID from the eastern part of India. Materials and Methods: Data from National Family Health Survey 4 were analyzed for three states - Bihar, West Bengal, and Jharkhand, keeping district as a unit of analysis. Correlation and regression were used for finding out the determinants of ID. Results: Overall, 67.8% of deliveries were conducted in hospitals. All the three states were able to improve performance on related health parameters like antenatal care. After adjusting for other variables, on multiple linear regression, female literacy was significantly related to ID. Conclusions: Social parameters need to be focused for encouraging ID.

Author Correction: Computationally designed antibody-drug conjugates self-assembled via affinity ligands.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

SNX27-retromer assembly recycles MT1-MMP to invadopodia and promotes breast cancer metastasis.

A variety of metastatic cancer cells use actin-rich membrane protrusions, known as invadopodia, for efficient ECM degradation, which involves trafficking of proteases from intracellular compartments to these structures. Here, we demonstrate that in the metastatic breast cancer cell line MDA-MB-231, retromer regulates the matrix invasion activity by recycling matrix metalloprotease, MT1-MMP. We further found that MT2-MMP, another abundantly expressed metalloprotease, is also invadopodia associated. MT1- and MT2-MMP showed a high degree of colocalization but were located on the distinct endosomal domains. Retromer and its associated sorting nexin, SNX27, phenocopied each other in matrix degradation via selectively recycling MT1-MMP but not MT2-MMP. ITC-based studies revealed that both SNX27 and retromer could directly interact with MT1-MMP. Analysis from a publicly available database showed SNX27 to be overexpressed or frequently altered in the patients having invasive breast cancer. In xenograft-based studies, SNX27-depleted cell lines showed prolonged survival of SCID mice, suggesting a possible implication for overexpression of the sorting nexin in tumor samples.

Computationally designed antibody-drug conjugates self-assembled via affinity ligands.

Antibody-drug conjugates (ADCs) combine the high specificity of antibodies with cytotoxic payloads. However, the present strategies for the synthesis of ADCs either yield unstable or heterogeneous products or involve complex processes. Here, we report a computational approach that leverages molecular docking and molecular dynamics simulations to design ADCs that self-assemble through the non-covalent binding of the antibody to a payload that we designed to act as an affinity ligand for specific conserved amino acid residues in the antibody. This method does not require modifications to the antibody structure and yields homogenous ADCs that form in less than 8 min. We show that two conjugates, which consist of hydrophilic and hydrophobic payloads conjugated to two different antibodies, retain the structure and binding properties of the antibody and its biological specificity, are stable in plasma and improve anti-tumour efficacy in mice with non-small cell lung tumour xenografts. The relative simplicity of the approach may facilitate the production of ADCs for the targeted delivery of cytotoxic payloads.

Enhanced Luminescent Properties of Photo-Stable Copper Nanoclusters through Formation of "Protein-Corona"-Like Assemblies.

In this study, interactions of synthesized copper nanoclusters (CuNCs) with a model transport protein, human serum albumin (HSA), have been systematically investigated by using various spectroscopic approaches. The interactions give rise to the formation of "protein-corona" like assemblies and the luminescence properties (both steady-state and time-resolved) are enhanced due to gradual adsorption of the protein on the surface of the NCs. The associated thermodynamics and binding parameters have been estimated resorting to luminescent experimental techniques as well as isothermal titration calorimetry (ITC) studies, indicating that every NC is surrounded by (4±1) protein molecules. The adsorption of HSA on the surface of the NCs has been characterized by dynamic light scattering (DLS) and time-resolved anisotropy measurements. Finally, fluorescence correlation spectroscopy (FCS) data substantiate the emergence of new "protein-corona" like assemblies resulting in slower translational diffusion motions and concomitant rise of the hydrodynamic diameters.

Structural and thermodynamic characterization of metal binding in Vps29 from Entamoeba histolytica: implication in retromer function.

Vps29 is the smallest subunit of retromer complex with metallo-phosphatase fold. Although the role of metal in Vps29 is in quest, its metal binding mutants has been reported to affect the localization of the retromer complex in human cells. In this study, we report the structural and thermodynamic consequences of these mutations in Vps29 from the protozoan parasite, Entamoeba histolytica (EhVps29). EhVps29 is a zinc binding protein as revealed by X-ray crystallography and isothermal titration calorimetry. The metal binding pocket of EhVps29 exhibits marked differences in its 3-dimensional architecture and metal coordination in comparison to its human homologs and other metallo-phosphatases. Alanine substitutions of the metal-coordinating residues showed significant alteration in the binding affinity of EhVps29 for zinc. We also determined the crystal structures of metal binding defective mutants (D62A and D62A/H86A) of EhVps29. Based on our results, we propose that the metal atoms or the bound water molecules in the metal binding site are important for maintaining the structural integrity of the protein. Further cellular studies in the amoebic trophozoites showed that the overexpression of wild type EhVps29 leads to reduction in intracellular cysteine protease activity suggesting its crucial role in secretion of the proteases.

Biophysical studies on calcium and carbohydrate binding to carbohydrate recognition domain of Gal/GalNAc lectin from Entamoeba histolytica: insights into host cell adhesion.

Entamoeba histolytica, an enteric parasite expresses a Gal/GalNAc-specific lectin that contributes to its virulence by establishing adhesion to host cell. In this study, carbohydrate recognition domain of Hgl (EhCRD) was purified and biophysical studies were conducted to understand the thermodynamic basis of its binding to carbohydrate and Ca(++) Here, we show that carbohydrate recognition domain (CRD) of the lectin binds to calcium through DPN motif. To decipher the role of calcium in carbohydrate binding and host cell adhesion, biophysical and cell-based studies were carried out. We demonstrated that the presence of the cation neither change the affinity of the lectin for carbohydrates nor alters its conformation. Mutation of the calcium-binding motif in EhCRD resulted in complete loss of ability to bind calcium but retained its affinity for carbohydrates. Purified EhCRD significantly diminished adhesion of the amebic trophozoites to Chinese Hamster Ovary (CHO) cells as well as triggered red blood cell agglutination. The calcium-binding defective mutant abrogated amebic adhesion to CHO cells similar to the wild-type protein, but it failed to agglutinate RBCs suggesting a differential role of the cation in these two processes. This study provides the first molecular description of the role of calcium in Gal/GalNAc mediated host cell adhesion.

Insights into the GTP/GDP cycle of RabX3, a novel GTPase from Entamoeba histolytica with tandem G-domains.

Members of the small GTPase Ras superfamily regulate a host of systems through their ability to catalyze the GTP/GDP cycle. All family members reported thus far possess a single GTPase domain with a P-loop containing a nucleoside triphosphate hydrolase fold. Here for the first time we report a novel member from Entamoeba histolytica, EhRabX3, which harbors two GTPase domains in tandem and exhibits unique biochemical properties. A combination of biochemical and microcalorimetric studies revealed that EhRabX3 binds to a single guanine nucleotide through its N-terminal domain. Unlike most of the members of the Ras superfamily, the dissociation of the nucleotide from EhRabX3 is independent of Mg(2+), perhaps indicating a novel mechanism of nucleotide exchange by this protein. We found that EhRabX3 is extremely sluggish in hydrolyzing GTP, and that could be attributed to its atypical nucleotide binding pocket. It harbors substitutions at two positions that confer oncogenicity to Ras because of impaired GTP hydrolysis. Engineering these residues into the conserved counterparts enhanced their GTPase activity by at least 20-fold. In contrast to most of the members of the Ras superfamily, EhRabX3 lacks the prenylation motif. Using indirect immunofluorescence and biochemical fractionation, we demonstrated that the protein is distributed all over the cytosol in amoebic trophozoites. Collectively, this unique ancient GTPase exhibits a striking evolutionary divergence from the other members of the superfamily.

Exploring the self-assembly of a short aromatic Aß(16-24) peptide.

The use of self-assembling peptides as scaffolds for creating biomaterials has prompted the scientific community to carry out studies on short peptides as model systems. Short peptides help in dissecting contributions from different interactions, unlike large peptides, where multiple interactions make it difficult to dissect the contributions of individual interactions. This opens avenues for fine tuning peptides to carry out a wide range of physical or chemical properties. In this line of study Aß(16-24) is a versatile building block not only as a scaffold for creating biomaterials but also because it forms the active core in the protein that forms amyloid plaques. In this study, we probe the self-assembly of peptide Aß(16-24) using fluorescence spectroscopy, circular dichroism, isothermal titration calorimetry, transmission electron microscopy, and atomic force microscopy. The process of self-assembly is dictated by the burial of phenyl alanines in the hydrophobic core and guided by nonbonding interactions and H-bonding. The process of fibril formation is enthalpically driven, and the fibrils showed blue and green luminescence without the addition of any external agent or sensitizer. Because these short peptides are known to bind with fully formed amyloid fibrils, this opens a route to the study of amyloid systems in vitro or isolated from patients suffering from Alzheimer's disease.

Metal-binding properties and structural characterization of a self-assembled coiled coil: formation of a polynuclear Cd-thiolate cluster.

This paper describes the design, characterization, and metal-binding properties of a 32-residue polypeptide called AQ-C16C19. The sequence of this peptide is composed of four repeats of the seven residue sequence Ile-Ala-Ala-Leu-Glu-Gln-Lys but with a Cys-X-X-Cys metal-binding motif substituted at positions 16-19. Size exclusion chromatography with multiangle light scattering detection (SEC-MALS) and circular dichroism (CD) spectroscopy studies showed that the apo peptide exhibits a pH-dependent oligomerization state in which a three-stranded α-helical coiled coil is dominant between pH5.4 and 8.5. The Cd(2+)-binding properties of the AQ-C16C19 peptide were studied by ultraviolet-visible spectroscopy (UV-vis), electrospray ionization mass spectrometry (ESI MS), and (113)Cd NMR techniques. The holoprotein was found to contain a polynuclear cadmium-thiolate center formed within the hydrophobic core of the triple-stranded α-helical coiled-coil structure. The X-ray crystal structure of the Cd-loaded peptide, resolved at 1.85Å resolution, revealed an adamantane-like configuration of the polynuclear metal center consisting of four cadmium ions, six thiolate sulfur ligands from cysteine residues and four oxygen-donor ligands. Three of these are from glutamic acid residues and one is from an exogenous water molecule. Thus, each cadmium ion is coordinated in a distorted tetrahedral S(3)O geometry. The metal cluster was found to form cooperatively at pH5.4 but in a stepwise fashion at pH>7. The results demonstrate that synthetic coiled-coils can be designed to incorporate multinuclear metal clusters, a proof-of-concept for their potential use in developing synthetic metalloenzymes and multi-electron redox agents.

Nanometer to millimeter scale peptide-porphyrin materials.

AQ-Pal14 is a 30-residue polypeptide that was designed to form an α-helical coiled coil that contains a metal-binding 4-pyridylalanine residue on its solvent-exposed surface. However, characterization of this peptide shows that it exists as a three-stranded coiled coil, not a two-stranded one as predicted from its design. Reaction with cobalt(III) protoporphyrin IX (Co-PPIX) produces a six-coordinate Co-PPIX(AQ-Pal14)(2) species that creates two coiled-coil oligomerization domains coordinated to opposite faces of the porphyrin ring. It is found that this species undergoes a buffer-dependent self-assembly process: nanometer-scale globular materials were formed when these components were reacted in unbuffered H(2)O, while millimeter-scale, rod-like materials were prepared when the reaction was performed in phosphate buffer (20 mM, pH 7). It is suggested that assembly of the globular material is dictated by the conformational properties of the coiled-coil forming AQ-Pal14 peptide, whereas that of the rod-like material involves interactions between Co-PPIX and phosphate ion.

Improvement of properties of polyetherimide/liquid crystalline polymer blends in the presence of functionalized carbon nanotubes.

This article reports the effect of functionalized multiwall carbon nanotubes (MWNT-COOH) on the morphological, dynamic mechanical, mechanical and thermal properties of polyetherimide (PEI)/liquid crystalline polymer (LCP) (Vectra A950) blends. The chemical modification of carbon nanotube enhanced the compatibility and the miscibility between PEI and LCP in the composites. Addition of functionalized MWNTs into the blend improved the thermal, mechanical and dynamic mechanical properties of the composite due to the presence of strong interfacial interaction between the polymer matrixes and the nanotubes in polymer composites. The glass transition temperature (from tan delta) increased from 208 degrees C to 245 degrees C with the addition of 1.8 wt% functionalized MWNTs in the blend of PEI/LCP. The tensile strength of the composite with 1.8 wt% MWNT-COOH was enhanced by 61% and 44% as compared to PEI/LCP blend and pure PEI. The functional groups on the MWNTs surface played an important role in accelerating both the dispersion of MWNTs and the interfacial adhesion in the composites compared to raw MWNTs.

Cd2+-induced conformational change of a synthetic metallopeptide: slow metal binding followed by a slower conformational change.

A two-stranded alpha-helical coiled coil was prepared having a Cys 4 metal-binding site within its hydrophobic interior. The addition of Cd2+ results in the incorporation of 2 equiv of metal ion, which is accompanied by a conformational change of the peptide, as observed by circular dichroism (CD) spectroscopy. Isothermal titration calorimetry (ITC) shows that the addition of Cd2+ is accompanied by two thermodynamic events. A comparison of the time dependence of the ITC behavior with those of the UV absorption and CD behavior allows the assignment of these events to a preliminary endothermic metal-binding step followed by a slower exothermic conformational change.