Hybrid-Integrated Biosensor for Express Determination of Protein Markers of Diseases based on Molecular Recognition and Direct Fluorimetric Detection

Ways of creating new generation biosensors for multiparametric express diagnostics based on molecular recognition and direct fluorimetric registration of a peptide aptamer — protein marker complex were considered. The biosensor platform comprises a microfluidic channel for delivery sample solutions, coupled with flow-through zones containing covalently attached arrays of peptide probes — aptamers. An outer glass window of the biochip assembly contains a layer of luminophore ZnS:Cu, bound on it via an acrylic lacquer and intended for the re-emitting native fluorescence of bound proteins into the longer wavelength range, more efficient in registering signals with CMOS sensors. The aptamers were designed using "Protein 3D" program for analysis of spatial complementarity of protein structures. The peptide, complementary to Troponin T, was modified by replacement of aromatic amino acid residue while maintaining the spatial configuration. The complementarity of peptide and Troponin T was confirmed using a capillary electrophoresis-on-a-chip. Biosensors are manufactured using thick-film technology and photolithography. The fluorescence of marker proteins was excited using UV-LED with a radiation wavelength of 275 nm. The limit of detection achieved for Troponin T was 6 ng/ml.

Unraveling the Compositional and Molecular Features Involved in Lysozyme-Benzothiazole Derivative Interactions

In this work we present a computational analysis together with experimental studies, focusing on the interaction between a benzothiazole (BTS) and lysozyme. Results obtained from isothermal titration calorimetry, UV-vis, and fluorescence were contrasted and complemented with molecular docking and machine learning techniques. The free energy values obtained both experimentally and theoretically showed excellent similarity. Calorimetry, UV-vis, and 3D/2D-lig-plot analysis revealed that the most relevant interactions between BTS and lysozyme are based on a predominance of aromatic, hydrophobic Van der Waals interactions, mainly aromatic edge-to-face (T-shaped) π-π stacking interactions between the benzene ring belonging to the 2-(methylthio)-benzothiazole moiety of BTS and the aromatic amino acid residue TRP108 of the lysozyme receptor. Next, conventional hydrogen bonding interactions contribute to the stability of the BTS-lysozyme coupling complex. In addition, mechanistic approaches performed using elastic network models revealed that the BTS ligand theoretically induces propagation of allosteric signals, suggesting non-physiological conformational flexing in large blocks of lysozyme affecting α-helices. Likewise, the BTS ligand interacts directly with allosteric residues, inducing perturbations in the conformational dynamics expressed as a moderate conformational softening in the α-helices H1, H2, and their corresponding β-loop in the lysozyme receptor, in contrast to the unbound state of lysozyme.

eMap: A Web Application for Identifying and Visualizing Electron or Hole Hopping Pathways in Proteins

<p>eMap is a web-based platform for identifying and visualizing electron or hole transfer pathways in proteins based on their crystal structures. The underlying model can be viewed as a coarse-grained version of the Pathways model, where each tunneling step between hopping sites represented by electron transfer active (ETA) moieties is described with one effective decay parameter that describes protein-mediated tunneling. ETA moieties include aromatic amino acid residue side chains and aromatic fragments of cofactors that are automatically detected, and, in addition, electron/hole residing sites that can be specified by the users. The software searches for the shortest paths connecting the user-specified electron/hole source to either all surface-exposed ETA residues or to the user-specified target. The identified pathways are ranked based on their length. The pathways are visualized in 2D as a graph, in which each node represents an ETA site, and in 3D using available protein visualization tools. Here, we present the capability and user interface of eMap 1.0, which is available at https://emap.bu.edu.</p>

eMap: A Web Application for Identifying and Visualizing Electron or Hole Hopping Pathways in Proteins

<p>eMap is a web-based platform for identifying and visualizing electron or hole transfer pathways in proteins based on their crystal structures. The underlying model can be viewed as a coarse-grained version of the Pathways model, where each tunneling step between hopping sites represented by electron transfer active (ETA) moieties is described with one effective decay parameter that describes protein-mediated tunneling. ETA moieties include aromatic amino acid residue side chains and aromatic fragments of cofactors that are automatically detected, and, in addition, electron/hole residing sites that can be specified by the users. The software searches for the shortest paths connecting the user-specified electron/hole source to either all surface-exposed ETA residues or to the user-specified target. The identified pathways are ranked based on their length. The pathways are visualized in 2D as a graph, in which each node represents an ETA site, and in 3D using available protein visualization tools. Here, we present the capability and user interface of eMap 1.0, which is available at https://emap.bu.edu.</p>

eMap: A Web Application for Identifying and Visualizing Electron or Hole Hopping Pathways in Proteins

<p>eMap is a web-based platform for identifying and visualizing electron or hole transfer pathways in proteins based on their crystal structures. The underlying model can be viewed as a coarse-grained version of the Pathways model with only through-space tunneling between electron transfer active (ETA) moieties being taken into account. ETA moieties include aromatic amino acid residue side chains and aromatic fragments of cofactors that are automatically detected, and, in addition, electron/hole residing sites that can be specified by the users. The software searches for the shortest paths connecting the user-specified electron/hole source to either all surface-exposed ETA residues or to the user-specified target. The identified pathways are ranked based on their length. The pathways are visualized in 2D as a graph, in which each node represents an ETA site, and in 3D using available protein visualization tools. Here, we present the capability and user interface of eMap 1.0, which is available at https://emap.bu.edu.</p>

eMap: a Web Application for Identifying and Visualizing Electron or Hole Hopping Pathways in Proteins

eMap is a web-based platform for identifying and visualizing electron or hole transfer pathways in proteins based on their crystal structures. The underlying model can be viewed as a coarse-grained version of the <i>Pathways</i> model with only through-space tunneling between electron transfer active (ETA) moieties being taken into account. ETA moieties include aromatic amino acid residue side chains and aromatic fragments of cofactors that are automatically detected, and, in addition, electron/hole residing sites that can be specified by the users. The software searches for the shortest paths connecting the user-specified electron/hole source to either all surface-exposed ETA residues or to the user-specified target. The identified pathways are ranked based on their length. The pathways are visualized in 2D as a graph, in which each node represents an ETA site, and in 3D using available protein visualization tools. Here we present the capability and user interface of eMap 1.0, which is available at https://emap.bu.edu.

Exploring cucurbit[6]uril–peptide interactions in the solid state: crystal structure of cucurbit[6]uril complexes with glycyl-containing dipeptides

Macrocyclic host cucurbit[6]uril forms supramolecular complexes with dipeptides sequenced as Gly-X, where X is either an aromatic amino acid residue Phe, Tyr, and Trp or Gly in the solid state.

Identification of Three Interferon-Inducible Cellular Enzymes That Inhibit the Replication of Hepatitis C Virus

ABSTRACT Hepatitis C virus (HCV) infection is a common cause of chronic hepatitis and is currently treated with alpha interferon (IFN-α)-based therapies. However, the underlying mechanism of IFN-α therapy remains to be elucidated. To identify the cellular proteins that mediate the antiviral effects of IFN-α, we created a HEK293-based cell culture system to inducibly express individual interferon-stimulated genes (ISGs) and determined their antiviral effects against HCV. By screening 29 ISGs that are induced in Huh7 cells by IFN-α and/or up-regulated in HCV-infected livers, we discovered that viperin, ISG20, and double-stranded RNA-dependent protein kinase (PKR) noncytolytically inhibited the replication of HCV replicons. Mechanistically, inhibition of HCV replication by ISG20 and PKR depends on their 3′-5′ exonuclease and protein kinase activities, respectively. Moreover, our work, for the first time, provides strong evidence suggesting that viperin is a putative radical S-adenosyl-l-methionine (SAM) enzyme. In addition to demonstrating that the antiviral activity of viperin depends on its radical SAM domain, which contains conserved motifs to coordinate [4Fe-4S] cluster and cofactor SAM and is essential for its enzymatic activity, mutagenesis studies also revealed that viperin requires an aromatic amino acid residue at its C terminus for proper antiviral function. Furthermore, although the N-terminal 70 amino acid residues of viperin are not absolutely required, deletion of this region significantly compromises its antiviral activity against HCV. Our findings suggest that viperin represents a novel antiviral pathway that works together with other antiviral proteins, such as ISG20 and PKR, to mediate the IFN response against HCV infection.