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Sample records for molecular recognition scaffolds

  1. Ion and Molecular Recognition Using Aryl–Ethynyl Scaffolding

    PubMed Central

    Vonnegut, Chris L.; Tresca, Blakely W.

    2015-01-01

    The aryl–ethynyl linkage has been extensively employed in the construction of hosts for a variety of guests. Uses range from ion detection (e.g., of metal cations in the environment or industrial waste and of anions prevalent in nature), to molecular mimics for biological systems, and to applications targeting future safety issues (such as CO2 capture and indicators for the manufacture of chemical weapons). This Focus Review examines the utilization of the aryl–ethynyl linkage in engineering host molecules for a variety of different guests, and how the alkyne unit plays an integral part as both a rigid scaffolding section in host geometry design as well as a linker to allow conjugative communication between discrete π-electron systems. PMID:25586943

  2. Ion and molecular recognition using aryl-ethynyl scaffolding.

    PubMed

    Vonnegut, Chris L; Tresca, Blakely W; Johnson, Darren W; Haley, Michael M

    2015-03-01

    The aryl-ethynyl linkage has been extensively employed in the construction of hosts for a variety of guests. Uses range from ion detection (e.g., of metal cations in the environment or industrial waste and of anions prevalent in nature), to molecular mimics for biological systems, and to applications targeting future safety issues (such as CO2 capture and indicators for the manufacture of chemical weapons). This Focus Review examines the utilization of the aryl-ethynyl linkage in engineering host molecules for a variety of different guests, and how the alkyne unit plays an integral part as both a rigid scaffolding section in host geometry design as well as a linker to allow conjugative communication between discrete π-electron systems. PMID:25586943

  3. Adhiron: a stable and versatile peptide display scaffold for molecular recognition applications

    PubMed Central

    Tiede, Christian; Tang, Anna A. S.; Deacon, Sarah E.; Mandal, Upasana; Nettleship, Joanne E.; Owen, Robin L.; George, Suja E.; Harrison, David J.; Owens, Raymond J.; Tomlinson, Darren C.; McPherson, Michael J.

    2014-01-01

    We have designed a novel non-antibody scaffold protein, termed Adhiron, based on a phytocystatin consensus sequence. The Adhiron scaffold shows high thermal stability (Tm ca. 101°C), and is expressed well in Escherichia coli. We have determined the X-ray crystal structure of the Adhiron scaffold to 1.75 Å resolution revealing a compact cystatin-like fold. We have constructed a phage-display library in this scaffold by insertion of two variable peptide regions. The library is of high quality and complexity comprising 1.3 × 1010 clones. To demonstrate library efficacy, we screened against the yeast Small Ubiquitin-like Modifier (SUMO). In selected clones, variable region 1 often contained sequences homologous to the known SUMO interactive motif (V/I-X-V/I-V/I). Four Adhirons were further characterised and displayed low nanomolar affinities and high specificity for yeast SUMO with essentially no cross-reactivity to human SUMO protein isoforms. We have identified binders against >100 target molecules to date including as examples, a fibroblast growth factor (FGF1), platelet endothelial cell adhesion molecule (PECAM-1; CD31), the SH2 domain Grb2 and a 12-aa peptide. Adhirons are highly stable and well expressed allowing highly specific binding reagents to be selected for use in molecular recognition applications. PMID:24668773

  4. Understanding selective molecular recognition in integrated carbon nanotube-polymer sensors by simulating physical analyte binding on carbon nanotube-polymer scaffolds.

    PubMed

    Lin, Shangchao; Zhang, Jingqing; Strano, Michael S; Blankschtein, Daniel

    2014-08-28

    Macromolecular scaffolds made of polymer-wrapped single-walled carbon nanotubes (SWCNTs) have been explored recently (Zhang et al., Nature Nanotechnology, 2013) as a new class of molecular-recognition motifs. However, selective analyte recognition is still challenging and lacks the underlying fundamental understanding needed for its practical implementation in biological sensors. In this report, we combine coarse-grained molecular dynamics (CGMD) simulations, physical adsorption/binding theories, and photoluminescence (PL) experiments to provide molecular insight into the selectivity of such sensors towards a large set of biologically important analytes. We find that the physical binding affinities of the analytes on a bare SWCNT partially correlate with their distribution coefficients in a bulk water/octanol system, suggesting that the analyte hydrophobicity plays a key role in determining the binding affinities of the analytes considered, along with the various specific interactions between the analytes and the polymer anchor groups. Two distinct categories of analytes are identified to demonstrate a complex picture for the correlation between optical sensor signals and the simulated binding affinities. Specifically, a good correlation was found between the sensor signals and the physical binding affinities of the three hormones (estradiol, melatonin, and thyroxine), the neurotransmitter (dopamine), and the vitamin (riboflavin) to the SWCNT-polymer scaffold. The four amino acids (aspartate, glycine, histidine, and tryptophan) and the two monosaccharides (fructose and glucose) considered were identified as blank analytes which are unable to induce sensor signals. The results indicate great success of our physical adsorption-based model in explaining the ranking in sensor selectivities. The combined framework presented here can be used to screen and select polymers that can potentially be used for creating synthetic molecular recognition motifs. PMID:24992310

  5. UFSRAT: Ultra-Fast Shape Recognition with Atom Types –The Discovery of Novel Bioactive Small Molecular Scaffolds for FKBP12 and 11βHSD1

    PubMed Central

    Shave, Steven; Blackburn, Elizabeth A.; Adie, Jillian; Houston, Douglas R.; Auer, Manfred; Webster, Scott P.; Taylor, Paul; Walkinshaw, Malcolm D.

    2015-01-01

    Motivation Using molecular similarity to discover bioactive small molecules with novel chemical scaffolds can be computationally demanding. We describe Ultra-fast Shape Recognition with Atom Types (UFSRAT), an efficient algorithm that considers both the 3D distribution (shape) and electrostatics of atoms to score and retrieve molecules capable of making similar interactions to those of the supplied query. Results Computational optimization and pre-calculation of molecular descriptors enables a query molecule to be run against a database containing 3.8 million molecules and results returned in under 10 seconds on modest hardware. UFSRAT has been used in pipelines to identify bioactive molecules for two clinically relevant drug targets; FK506-Binding Protein 12 and 11β-hydroxysteroid dehydrogenase type 1. In the case of FK506-Binding Protein 12, UFSRAT was used as the first step in a structure-based virtual screening pipeline, yielding many actives, of which the most active shows a KD, app of 281 µM and contains a substructure present in the query compound. Success was also achieved running solely the UFSRAT technique to identify new actives for 11β-hydroxysteroid dehydrogenase type 1, for which the most active displays an IC50 of 67 nM in a cell based assay and contains a substructure radically different to the query. This demonstrates the valuable ability of the UFSRAT algorithm to perform scaffold hops. Availability and Implementation A web-based implementation of the algorithm is freely available at http://opus.bch.ed.ac.uk/ufsrat/. PMID:25659145

  6. Bioimprinted Polymer Scaffolds for Selective Recognition of RGD Peptides

    NASA Astrophysics Data System (ADS)

    Bergmann, Nicole; Peppas, Nicholas A.

    2003-03-01

    Fibronectin and a number of other plasma and extracellular matrix (ECM) adhesion proteins contain the tetrapeptide arginine-glycine-aspartic acid-serine (RGDS), and this sequence can be summarily recognized and bound by integrins present on cell membranes. Upon integrin binding, cells adhere to the substrate, and this adherence encourages ECM deposition and other cellular remodeling events. By targeting specific chemical functional groups on the peptide using non-covalent molecular imprinting, biomimetic polymeric scaffolds can be designed to mimic protein-ECM binding both on the surface and in the bulk during polymer degradation. Methacrylic acid-ethylene glycol dimethacrylate (MAA-g-EGDMA) copolymer films were prepared by free-radical ultraviolet polymerization in the presence of RGDS to create novel imprinted matrices for possible tissue engineering scaffolds. SEM analysis revealed a highly macroporous structure in peptide-imprinted polymers compared to controls. Optimal crosslinking ratios for peptide imprinting were determined using a small molecular weight fluorescent tag, 4-chloro-7-nitrobenzofurazan, and analyzed using fluorescent microscopy. Higher crosslinking ratios yielded better template recognition and gels exhibited specific recognition in aqueous media to RGDS molecules when in the presence of similar tetrapeptides.

  7. Scaffolding Learning from Molecular Visualizations

    ERIC Educational Resources Information Center

    Chang, Hsin-Yi; Linn, Marcia C.

    2013-01-01

    Powerful online visualizations can make unobservable scientific phenomena visible and improve student understanding. Instead, they often confuse or mislead students. To clarify the impact of molecular visualizations for middle school students we explored three design variations implemented in a Web-based Inquiry Science Environment (WISE) unit on…

  8. Molecular Recognition and Ligand Association

    NASA Astrophysics Data System (ADS)

    Baron, Riccardo; McCammon, J. Andrew

    2013-04-01

    We review recent developments in our understanding of molecular recognition and ligand association, focusing on two major viewpoints: (a) studies that highlight new physical insight into the molecular recognition process and the driving forces determining thermodynamic signatures of binding and (b) recent methodological advances in applications to protein-ligand binding. In particular, we highlight the challenges posed by compensating enthalpic and entropic terms, competing solute and solvent contributions, and the relevance of complex configurational ensembles comprising multiple protein, ligand, and solvent intermediate states. As more complete physics is taken into account, computational approaches increase their ability to complement experimental measurements, by providing a microscopic, dynamic view of ensemble-averaged experimental observables. Physics-based approaches are increasingly expanding their power in pharmacology applications.

  9. Muscle Giants: Molecular Scaffolds in Sarcomerogenesis

    PubMed Central

    KONTROGIANNI-KONSTANTOPOULOS, AIKATERINI; ACKERMANN, MAEGEN A.; BOWMAN, AMBER L.; YAP, SOLOMON V.; BLOCH, ROBERT J.

    2011-01-01

    Myofibrillogenesis in striated muscles is a highly complex process that depends on the coordinated assembly and integration of a large number of contractile, cytoskeletal, and signaling proteins into regular arrays, the sarcomeres. It is also associated with the stereotypical assembly of the sarcoplasmic reticulum and the transverse tubules around each sarcomere. Three giant, muscle-specific proteins, titin (3–4 MDa), nebulin (600–800 kDa), and obscurin (~720–900 kDa), have been proposed to play important roles in the assembly and stabilization of sarcomeres. There is a large amount of data showing that each of these molecules interacts with several to many different protein ligands, regulating their activity and localizing them to particular sites within or surrounding sarcomeres. Consistent with this, mutations in each of these proteins have been linked to skeletal and cardiac myopathies or to muscular dystrophies. The evidence that any of them plays a role as a “molecular template,” “molecular blueprint,” or “molecular ruler” is less definitive, however. Here we review the structure and function of titin, nebulin, and obscurin, with the literature supporting a role for them as scaffolding molecules and the contradictory evidence regarding their roles as molecular guides in sarcomerogenesis. PMID:19789381

  10. Characterization of molecular recognition in gas sensors

    SciTech Connect

    Hierlemann, A.; Ricco, A.J.; Bodenhoefer, K.; Goepel, W.

    1998-08-01

    Molecular recognition is an important topic when searching for new, selective coating materials for chemical sensing. Recently, the general idea of molecular recognition in the gas phase was challenged by Grate et al. However, in earlier thickness-shear mode resonator (TSMR) investigations, convincing evidence was presented for specific recognition of particular analyte target molecules. In this study, the authors systematically investigated coatings previously shown to be highly selective, such as the bucket-like cyclodextrins for chiral recognition, Ni-camphorates for the specific detection of the bases pyridine and DMMP (dimethylmethylphosphonate), and phthalocyanines to specifically detect benzene, toluene, and xylene (BTX).

  11. Molecular Recognition of Natural Products by Resorc[4]arene Receptors.

    PubMed

    D'Acquarica, Ilaria; Ghirga, Francesca; Quaglio, Deborah; Cerreto, Antonella; Ingallina, Cinzia; Tafi, Andrea; Botta, Bruno

    2016-01-01

    This review is aimed at providing an overview of the up-to-now published literature on resorc[4]arene macrocycles exploited as artificial receptors for the molecular recognition of some classes of natural products. A concise illustration of the main synthetic strategies developed to afford the resorc[4]arene scaffold is followed by a report on the principles of the gas-phase investigation of recognition phenomena by mass spectrometry (MS). Emphasis is placed on gas-phase studies of diastereoisomeric complexes generated inside a Fourier transform-ion cyclotron resonance (FT-ICR) mass spectrometer by resorc[4]arene receptors towards a series of natural products, namely amino acids, amphetamine, ethanolamine neurotransmitters, dipeptides, vinca alkaloids and nucleosides. The literature outcomes discussed here, taken largely from our own revisited work, have been completed by references to other studies, in order to draw a broader picture of this rapidly evolving field of research. PMID:26654589

  12. Plastic Antibodies: Molecular Recognition with Imprinted Polymers

    ERIC Educational Resources Information Center

    Rushton, Gregory T.; Furmanski, Brian; Shimizu, Ken D.

    2005-01-01

    Synthetic polymers are prepared and tested in a study for their molecular recognition properties of an adenine derivative, ethyl adenine-9-acetate (EA9A), within two laboratory periods. The procedure introduces undergraduate chemistry students to noncovalent molecular imprinting as well as the analytical techniques for assessing their recognition…

  13. Chalcone Scaffold in Anticancer Armamentarium: A Molecular Insight

    PubMed Central

    Manna, Kuntal

    2016-01-01

    Cancer is an inevitable matter of concern in the medicinal chemistry era. Chalcone is the well exploited scaffold in the anticancer domain. The molecular mechanism of chalcone at cellular level was explored in past decades. This mini review provides the most recent updates on anticancer potential of chalcones. PMID:26880913

  14. Molecular recognition: The I's have it

    NASA Astrophysics Data System (ADS)

    Taylor, Mark S.

    2014-12-01

    Rotaxanes with cyclodextrin end groups have been used as a platform to investigate anion binding in water, revealing that halogen bonding can serve as the basis for molecular recognition in aqueous solvents, which may have implications in medicinal chemistry and beyond.

  15. Studies on molecular recognition of thymidines with molecularly imprinted polymers

    NASA Astrophysics Data System (ADS)

    Chen, Zhen-He; Luo, Ai-Qin; Sun, Li-Quan

    2009-07-01

    Molecularly imprinted polymers (MIPs) with excellent molecular recognition ability have been used in chemical sensors, chromatographic separation and biochemical analyses. Thymidine is an important part of DNA for biomolecular recognition and the intermediate of many medicines. The polymers imprinted with the template of thymidine and 5'-Otosylthymidine have been prepared, using a non-proton solvent, acetonitrile as the porogen. Direct imprinting with thymidine could not form strong molecular interaction sites in this system. Relative MIPs were obtained by bulk polymerization and their adsorption capacities were investigated. The adsorption capacities of MIP (P2) and nonimprinted polymer (P20) for thymidine are 0.120 mg•g-1and 0.103 mg•g-1, respectively. The imprinting factor is 1.17. As 5'-O-tosylthymidine is more soluble than thymidine moiety in acetonitrile and give rise to more sites of molecular recognition. The results demonstrated that the imprinted polymers were able to bind and recognize thymidine moderately in acetonitrile. MIPs imprinted with 5'-O-tosylthymidine like nature enzymes displayed some recognition ability to its analogues. The insoluble derivatives in the non-proton solvent can be an effective template to prepare efficient imprinting recognition sites.

  16. Molecular Recognition of Lys and Arg Methylation.

    PubMed

    Beaver, Joshua E; Waters, Marcey L

    2016-03-18

    A network of reader proteins and enzymes precisely controls gene transcription through the dynamic addition, removal, and recognition of post-translational modifications (PTMs) of histone tails. Histone PTMs work in concert with this network to regulate gene transcription through the histone code, and the dysregulation of PTM maintenance is linked to a large number of diseases, including many types of cancer. A wealth of research aims to elucidate the functions of this code, but our understanding of the effects of PTMs, specifically the methylation of lysine (Lys) and arginine (Arg), is lacking. The development of new tools to study PTMs relies on a sophisticated understanding of the mechanisms that drive protein and small molecule recognition in water. In this review, we outline the physical organic concepts that drive the molecular recognition of Lys and Arg methylation by reader proteins and draw comparisons to the binding mechanisms of small molecule receptors for methylated Lys and Arg that have been developed recently. PMID:26759915

  17. Photoswitchable gel assembly based on molecular recognition.

    PubMed

    Yamaguchi, Hiroyasu; Kobayashi, Yuichiro; Kobayashi, Ryosuke; Takashima, Yoshinori; Hashidzume, Akihito; Harada, Akira

    2012-01-01

    The formation of effective and precise linkages in bottom-up or top-down processes is important for the development of self-assembled materials. Self-assembly through molecular recognition events is a powerful tool for producing functionalized materials. Photoresponsive molecular recognition systems can permit the creation of photoregulated self-assembled macroscopic objects. Here we demonstrate that macroscopic gel assembly can be highly regulated through photoisomerization of an azobenzene moiety that interacts differently with two host molecules. A photoregulated gel assembly system is developed using polyacrylamide-based hydrogels functionalized with azobenzene (guest) or cyclodextrin (host) moieties. Reversible adhesion and dissociation of the host gel from the guest gel may be controlled by photoirradiation. The differential affinities of α-cyclodextrin or β-cyclodextrin for the trans-azobenzene and cis-azobenzene are employed in the construction of a photoswitchable gel assembly system. PMID:22215078

  18. Photoswitchable gel assembly based on molecular recognition

    PubMed Central

    Yamaguchi, Hiroyasu; Kobayashi, Yuichiro; Kobayashi, Ryosuke; Takashima, Yoshinori; Hashidzume, Akihito; Harada, Akira

    2012-01-01

    The formation of effective and precise linkages in bottom-up or top-down processes is important for the development of self-assembled materials. Self-assembly through molecular recognition events is a powerful tool for producing functionalized materials. Photoresponsive molecular recognition systems can permit the creation of photoregulated self-assembled macroscopic objects. Here we demonstrate that macroscopic gel assembly can be highly regulated through photoisomerization of an azobenzene moiety that interacts differently with two host molecules. A photoregulated gel assembly system is developed using polyacrylamide-based hydrogels functionalized with azobenzene (guest) or cyclodextrin (host) moieties. Reversible adhesion and dissociation of the host gel from the guest gel may be controlled by photoirradiation. The differential affinities of α-cyclodextrin or β-cyclodextrin for the trans-azobenzene and cis-azobenzene are employed in the construction of a photoswitchable gel assembly system. PMID:22215078

  19. Supramolecular Polymerization Engineered with Molecular Recognition.

    PubMed

    Haino, Takeharu

    2015-10-01

    Supramolecular polymeric assemblies represent an emerging, promising class of molecular assemblies with enormous versatility compared with their covalent polymeric counterparts. Although a large number of host-guest motifs have been produced over the history of supramolecular chemistry, only a limited number of recognition motifs have been utilized as supramolecular connections in polymeric assemblies. This account describes the molecular recognition of host molecules based on calix[5]arene and bisporphyrin that demonstrate unique guest encapsulations; subsequently, these host-guest motifs are applied to the synthesis of supramolecular polymers that display polymer-like properties in solution and solid states. In addition, new bisresorcinarenes are developed to form supramolecular polymers that are connected via a rim-to-rim hydrogen-bonded dimeric structure, which is composed of two resorcinarene moieties. PMID:26178364

  20. Molecular recognition in gels, monolayers, and solids

    NASA Astrophysics Data System (ADS)

    Prime, Kevin L.; Chu, Yen-Ho; Schmid, Walther; Seto, Christopher T.; Chen, James K.

    1991-12-01

    This paper describes work in four areas: affinity electrophoresis of carbonic anhydrase in cross-linked polyacrylamide derived gels containing immobilized derivatives of aryl sulfonamides; inhibition of the hemagglutination of erythrocytes induced by influenza virus using water-soluble polyacrylamides bearing sialic acid groups; the application of self-assembled monolayers (SAMs) of alkyl thiolates on gold to the study of protein adsorption on organic surfaces; and the use of networks of hydrogen bonds to generate new classes of non-covalently assembled organic materials, both in solution and in crystals. This paper summarizes research in two areas of molecular recognition: affinity polymers and molecular self assembly. We illustrate these areas by examples drawn from affinity gel electrophoresis, soluble synthetic macromolecular inhibitors of binding of influenza virus to erythrocytes protein adsorption on self assembled monolayers and self assembling hydrogen bonded molecular aggregates.

  1. Protein-targeted corona phase molecular recognition

    PubMed Central

    Bisker, Gili; Dong, Juyao; Park, Hoyoung D.; Iverson, Nicole M.; Ahn, Jiyoung; Nelson, Justin T.; Landry, Markita P.; Kruss, Sebastian; Strano, Michael S.

    2016-01-01

    Corona phase molecular recognition (CoPhMoRe) uses a heteropolymer adsorbed onto and templated by a nanoparticle surface to recognize a specific target analyte. This method has not yet been extended to macromolecular analytes, including proteins. Herein we develop a variant of a CoPhMoRe screening procedure of single-walled carbon nanotubes (SWCNT) and use it against a panel of human blood proteins, revealing a specific corona phase that recognizes fibrinogen with high selectivity. In response to fibrinogen binding, SWCNT fluorescence decreases by >80% at saturation. Sequential binding of the three fibrinogen nodules is suggested by selective fluorescence quenching by isolated sub-domains and validated by the quenching kinetics. The fibrinogen recognition also occurs in serum environment, at the clinically relevant fibrinogen concentrations in the human blood. These results open new avenues for synthetic, non-biological antibody analogues that recognize biological macromolecules, and hold great promise for medical and clinical applications. PMID:26742890

  2. Protein-targeted corona phase molecular recognition

    NASA Astrophysics Data System (ADS)

    Bisker, Gili; Dong, Juyao; Park, Hoyoung D.; Iverson, Nicole M.; Ahn, Jiyoung; Nelson, Justin T.; Landry, Markita P.; Kruss, Sebastian; Strano, Michael S.

    2016-01-01

    Corona phase molecular recognition (CoPhMoRe) uses a heteropolymer adsorbed onto and templated by a nanoparticle surface to recognize a specific target analyte. This method has not yet been extended to macromolecular analytes, including proteins. Herein we develop a variant of a CoPhMoRe screening procedure of single-walled carbon nanotubes (SWCNT) and use it against a panel of human blood proteins, revealing a specific corona phase that recognizes fibrinogen with high selectivity. In response to fibrinogen binding, SWCNT fluorescence decreases by >80% at saturation. Sequential binding of the three fibrinogen nodules is suggested by selective fluorescence quenching by isolated sub-domains and validated by the quenching kinetics. The fibrinogen recognition also occurs in serum environment, at the clinically relevant fibrinogen concentrations in the human blood. These results open new avenues for synthetic, non-biological antibody analogues that recognize biological macromolecules, and hold great promise for medical and clinical applications.

  3. Carbon nanodots as molecular scaffolds for development of antimicrobial agents.

    PubMed

    Ngu-Schwemlein, Maria; Chin, Suk Fun; Hileman, Ryan; Drozdowski, Chris; Upchurch, Clint; Hargrove, April

    2016-04-01

    We report the potential of carbon nanodots (CNDs) as a molecular scaffold for enhancing the antimicrobial activities of small dendritic poly(amidoamines) (PAMAM). Carbon nanodots prepared from sago starch are readily functionalized with PAMAM by using N-ethyl-N'-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDC) and N-hydroxysuccinimide (NHS). Electron microscopy images of these polyaminated CNDs show that they are approximately 30-60nm in diameter. Infrared and fluorescence spectroscopy analyses of the water-soluble material established the presence of the polyamidoaminated moiety and the intrinsic fluorescence of the nanodots. The polyaminated nanodots (CND-PAM1 and CND-PAM2) exhibit in vitro antimicrobial properties, not only to non-multidrug resistant bacteria but also to the corresponding Gram-negative multidrug bacteria. Their minimum inhibitory concentration (MIC) ranges from 8 to 64μg/mL, which is much lower than that of PAMAM G1 or the non-active PAMAM G0 and CNDs. Additionally, they show synergistic effect in combination with tetracycline or colistin. These preliminary results imply that CNDs can serve as a promising scaffold for facilitating the rational design of antimicrobial materials for combating the ever-increasing threat of antibiotic resistance. Moreover, their fluorescence could be pertinent to unraveling their mode of action for imaging or diagnostic applications. PMID:26923697

  4. Developments in Molecular Recognition and Sensing at Interfaces

    PubMed Central

    Ariga, Katsuhiko; Hill, Jonathan P.; Endo, Hiroshi

    2007-01-01

    In biological systems, molecular recognition events occur mostly within interfacial environments such as at membrane surfaces, enzyme reaction sites, or at the interior of the DNA double helix. Investigation of molecular recognition at model interfaces provides great insights into biological phenomena. Molecular recognition at interfaces not only has relevance to biological systems but is also important for modern applications such as high sensitivity sensors. Selective binding of guest molecules in solution to host molecules located at solid surfaces is crucial for electronic or photonic detection of analyte substances. In response to these demands, molecular recognition at interfaces has been investigated extensively during the past two decades using Langmuir monolayers, self-assembled monolayers, and lipid assemblies as recognition media. In this review, advances of molecular recognition at interfaces are briefly summarized.

  5. Molecular Recognition in the Digital Radio Domain

    NASA Astrophysics Data System (ADS)

    Hunt, William D.; Edmonson, Peter J.; Stubbs, Desmond D.; Lee, Sang-Hun

    2010-07-01

    In this paper we discuss the theoretical and experimental constructs which together point the way towards the transduction of biomolecular recognition events into a palpable set of electrical signals. This combines the applied physics of surface perturbations on acoustic wave device surfaces and the biochemistry of the interactions between an immobilized biomolecule (e.g., an antibody) and a target molecule which is flowing past the sensor surface (e.g., an antigen). We will first provide the theoretical basis for our contention that we can extract information about both molecular recognition and conformational change from the electrical signal and will then confirm this assertion with experimental results relating to induced conformational changes in DNA on a quartz crystal microbalance (QCM) surface. Next we will discuss our digital radio technique whereby the real time measurements using antibody coated surface acoustic wave (SAW) devices in the vapor phase allow us to differentiate between close chemical analogs of nitro-based molecules (e.g., tri-nitro toluene vs musk oil) by virtue of the cross-reactivity of the antibody-antigen interaction. In immunochemistry this is referred to as antibody promiscuity. Finally, we present two- and three-dimensional plots illustrating our technique which derives much from in-phase and quadrature phase (IQ) mapping. The end result is a powerful technique which allows one to differentiate between target molecules and chemically similar interferrents.

  6. Molecular recognition in chemical and biological systems.

    PubMed

    Persch, Elke; Dumele, Oliver; Diederich, François

    2015-03-01

    Structure-based ligand design in medicinal chemistry and crop protection relies on the identification and quantification of weak noncovalent interactions and understanding the role of water. Small-molecule and protein structural database searches are important tools to retrieve existing knowledge. Thermodynamic profiling, combined with X-ray structural and computational studies, is the key to elucidate the energetics of the replacement of water by ligands. Biological receptor sites vary greatly in shape, conformational dynamics, and polarity, and require different ligand-design strategies, as shown for various case studies. Interactions between dipoles have become a central theme of molecular recognition. Orthogonal interactions, halogen bonding, and amide⋅⋅⋅π stacking provide new tools for innovative lead optimization. The combination of synthetic models and biological complexation studies is required to gather reliable information on weak noncovalent interactions and the role of water. PMID:25630692

  7. Molecular Recognition and Specific Interactions for Biosensing Applications

    PubMed Central

    Kim, Dong Chung; Kang, Dae Joon

    2008-01-01

    Molecular recognition and specific interactions are reliable and versatile routes for site-specific and well-oriented immobilization of functional biomolecules on surfaces. The control of surface properties via the molecular recognition and specific interactions at the nanoscale is a key element for the nanofabrication of biosensors with high sensitivity and specificity. This review intends to provide a comprehensive understanding of the molecular recognition- and specific interaction-mediated biosensor fabrication routes that leads to biosensors with well-ordered and controlled structures on both nanopatterned surfaces and nanomaterials. Herein self-assembly of the biomolecules via the molecular recognition and specific interactions on nanoscaled surfaces as well as nanofabrication techniques of the biomolecules for biosensor architecture are discussed. We also describe the detection of molecular recognition- and specific interaction-mediated molecular binding as well as advantages of nanoscale detection.

  8. Molecular Recognition: Detection of Colorless Compounds Based on Color Change

    ERIC Educational Resources Information Center

    Khalafi, Lida; Kashani, Samira; Karimi, Javad

    2016-01-01

    A laboratory experiment is described in which students measure the amount of cetirizine in allergy-treatment tablets based on molecular recognition. The basis of recognition is competition of cetirizine with phenolphthalein to form an inclusion complex with ß-cyclodextrin. Phenolphthalein is pinkish under basic condition, whereas it's complex form…

  9. Molecular recognition by gold, silver and copper nanoparticles

    PubMed Central

    Tauran, Yannick; Brioude, Arnaud; Coleman, Anthony W; Rhimi, Moez; Kim, Beonjoom

    2013-01-01

    The intrinsic physical properties of the noble metal nanoparticles, which are highly sensitive to the nature of their local molecular environment, make such systems ideal for the detection of molecular recognition events. The current review describes the state of the art concerning molecular recognition of Noble metal nanoparticles. In the first part the preparation of such nanoparticles is discussed along with methods of capping and stabilization. A brief discussion of the three common methods of functionalization: Electrostatic adsorption; Chemisorption; Affinity-based coordination is given. In the second section a discussion of the optical and electrical properties of nanoparticles is given to aid the reader in understanding the use of such properties in molecular recognition. In the main section the various types of capping agents for molecular recognition; nucleic acid coatings, protein coatings and molecules from the family of supramolecular chemistry are described along with their numerous applications. Emphasis for the nucleic acids is on complementary oligonucleotide and aptamer recognition. For the proteins the recognition properties of antibodies form the core of the section. With respect to the supramolecular systems the cyclodextrins, calix[n]arenes, dendrimers, crown ethers and the cucurbitales are treated in depth. Finally a short section deals with the possible toxicity of the nanoparticles, a concern in public health. PMID:23977421

  10. Polymer side-chains as arms for molecular recognition

    NASA Astrophysics Data System (ADS)

    South, Clinton Ray

    This thesis describes research based on synthetic protocols, methodologies, and applications of polymers containing side-chain molecular recognition elements. The motivation for the thesis lies in the belief among many in the field that a strict covalent paradigm for polymer chemistry is reaching its limit. The use of molecular recognition, in lieu of covalent chemistry, potentially presents a path through the current limits of polymer science. The work described in the following chapters of this thesis is, at least in part, a testament to this proposal. The first two chapters present a basic introduction and survey of the fundamental noncovalent interactions that are ubiquitous in the research of supramolecular polymers and molecular recognition. A hierarchy of noncovalent interactions, molecular recognition, and self-assembly is outlined and discussed. Chapter 2 lays the foundation for the remaining chapters of this thesis by presenting several examples of prior work related specifically to the use of molecular recognition on the side-chains of polymers. The next two chapters present research focused on advancing the functionalization of polymers through molecular recognition. The goal of this research is primarily to develop a general polymer backbone that both site-specifically and strongly associates noncovalently with small molecular substrates. These chapters demonstrate that both architecturally controlled block copolymers and random terpolymers can accept a full load of different substrates without interference among distinct molecular recognition elements along the polymer backbone. Chapters 5 and 6 present a unique application of polymers containing molecular recognition elements, templated synthesis. Chapter 5 first discusses lessons learned from small molecule based templated synthesis in which a template and a substrate are held together by metal coordination and a subsequent bond forming reaction occurs. Ultimately, the results of this chapter

  11. Supramolecular polymers constructed by crown ether-based molecular recognition.

    PubMed

    Zheng, Bo; Wang, Feng; Dong, Shengyi; Huang, Feihe

    2012-03-01

    Supramolecular polymers, polymeric systems beyond the molecule, have attracted more and more attention from scientists due to their applications in various fields, including stimuli-responsive materials, healable materials, and drug delivery. Due to their good selectivity and convenient enviro-responsiveness, crown ether-based molecular recognition motifs have been actively employed to fabricate supramolecular polymers with interesting properties and novel applications in recent years. In this tutorial review, we classify supramolecular polymers based on their differences in topology and cover recent advances in the marriage between crown ether-based molecular recognition and polymer science. PMID:22012256

  12. Creating molecular macrocycles for anion recognition

    PubMed Central

    2016-01-01

    Summary The creation and functionality of new classes of macrocycles that are shape persistent and can bind anions is described. The genesis of triazolophane macrocycles emerges out of activity surrounding 1,2,3-triazoles made using click chemistry; and the same triazoles are responsible for anion capture. Mistakes made and lessons learnt in anion recognition provide deeper understanding that, together with theory, now provides for computer-aided receptor design. The lessons are acted upon in the creation of two new macrocycles. First, cyanostars are larger and like to capture large anions. Second is tricarb, which also favors large anions but shows a propensity to self-assemble in an orderly and stable manner, laying a foundation for future designs of hierarchical nanostructures. PMID:27340452

  13. Molecular Handshake: Recognition through Weak Noncovalent Interactions

    ERIC Educational Resources Information Center

    Murthy, Parvathi S.

    2006-01-01

    The weak noncovalent interactions between substances, the handshake in the form of electrostatic interactions, van der Waals' interactions or hydrogen bonding is universal to all living and nonliving matter. They significantly influence the molecular and bulk properties and behavior of matter. Their transient nature affects chemical reactions and…

  14. FINAL REPORT. SENSORS USING MOLECULAR RECOGNITION IN LUMINESCENT, CONDUCTIVE POLYMERS

    EPA Science Inventory

    The purpose of this project is to develop sensor technology for detecting specific heavy metal ions, such as transition metals, lead, lanthanides, and actinides in waste streams. The sensing strategy uses molecular recognition of the metal ions by polymers that change their lumin...

  15. Modulating mechanical behaviour of 3D-printed cartilage-mimetic PCL scaffolds: influence of molecular weight and pore geometry.

    PubMed

    Olubamiji, Adeola D; Izadifar, Zohreh; Si, Jennifer L; Cooper, David M L; Eames, B Frank; Chen, Daniel X B

    2016-06-01

    Three-dimensional (3D)-printed poly(ε)-caprolactone (PCL)-based scaffolds are increasingly being explored for cartilage tissue engineering (CTE) applications. However, ensuring that the mechanical properties of these PCL-based constructs are comparable to that of articular cartilage that they are meant to regenerate is an area that has been under-explored. This paper presents the effects of PCL's molecular weight (MW) and scaffold's pore geometric configurations; strand size (SZ), strand spacing (SS), and strand orientation (SO), on mechanical properties of 3D-printed PCL scaffolds. The results illustrate that MW has significant effect on compressive moduli and yield strength of 3D-printed PCL scaffolds. Specifically, PCL with MW of 45 K was a more feasible choice for fabrication of visco-elastic, flexible and load-bearing PCL scaffolds. Furthermore, pore geometric configurations; SZ, SS, and SO, all significantly affect on tensile moduli of scaffolds. However, only SZ and SS have statistically significant effects on compressive moduli and porosity of these scaffolds. That said, inverse linear relationship was observed between porosity and mechanical properties of 3D-printed PCL scaffolds in Pearson's correlation test. Altogether, this study illustrates that modulating MW of PCL and pore geometrical configurations of the scaffolds enabled design and fabrication of PCL scaffolds with mechanical and biomimetic properties that better mimic mechanical behaviour of human articular cartilage. Thus, the modulated PCL scaffold proposed in this study is a framework that offers great potentials for CTE applications. PMID:27328736

  16. Molecular recognition in protein modification with rhodium metallopeptides

    PubMed Central

    Ball, Zachary T.

    2015-01-01

    Chemical manipulation of natural, unengineered proteins is a daunting challenge which tests the limits of reaction design. By combining transition-metal or other catalysts with molecular recognition ideas, it is possible to achieve site-selective protein reactivity without the need for engineered recognition sequences or reactive sites. Some recent examples in this area have used ruthenium photocatalysis, pyridine organocatalysis, and rhodium(II) metallocarbene catalysis, indicating that the fundamental ideas provide opportunities for using diverse reactivity on complex protein substrates and in complex cell-like environments. PMID:25588960

  17. Recent Progress in Molecular Recognition Imaging Using Atomic Force Microscopy.

    PubMed

    Senapati, Subhadip; Lindsay, Stuart

    2016-03-15

    Atomic force microscopy (AFM) is an extremely powerful tool in the field of bionanotechnology because of its ability to image single molecules and make measurements of molecular interaction forces with piconewton sensitivity. It works in aqueous media, enabling studies of molecular phenomenon taking place under physiological conditions. Samples can be imaged in their near-native state without any further modifications such as staining or tagging. The combination of AFM imaging with the force measurement added a new feature to the AFM technique, that is, molecular recognition imaging. Molecular recognition imaging enables mapping of specific interactions between two molecules (one attached to the AFM tip and the other to the imaging substrate) by generating simultaneous topography and recognition images (TREC). Since its discovery, the recognition imaging technique has been successfully applied to different systems such as antibody-protein, aptamer-protein, peptide-protein, chromatin, antigen-antibody, cells, and so forth. Because the technique is based on specific binding between the ligand and receptor, it has the ability to detect a particular protein in a mixture of proteins or monitor a biological phenomenon in the native physiological state. One key step for recognition imaging technique is the functionalization of the AFM tips (generally, silicon, silicon nitrides, gold, etc.). Several different functionalization methods have been reported in the literature depending on the molecules of interest and the material of the tip. Polyethylene glycol is routinely used to provide flexibility needed for proper binding as a part of the linker that carries the affinity molecule. Recently, a heterofunctional triarm linker has been synthesized and successfully attached with two different affinity molecules. This novel linker, when attached to AFM tip, helped to detect two different proteins simultaneously from a mixture of proteins using a so-called "two

  18. Characterization of monobody scaffold interactions with ligand via force spectroscopy and steered molecular dynamics.

    PubMed

    Cheung, Luthur Siu-Lun; Shea, Daniel J; Nicholes, Nathan; Date, Amol; Ostermeier, Marc; Konstantopoulos, Konstantinos

    2015-01-01

    Monobodies are antibody alternatives derived from fibronectin that are thermodynamically stable, small in size, and can be produced in bacterial systems. Monobodies have been engineered to bind a wide variety of target proteins with high affinity and specificity. Using alanine-scanning mutagenesis simulations, we identified two scaffold residues that are critical to the binding interaction between the monobody YS1 and its ligand, maltose-binding protein (MBP). Steered molecular dynamics (SMD) simulations predicted that the E47A and R33A mutations in the YS1 scaffold substantially destabilize the YS1-MBP interface by reducing the bond rupture force and the lifetime of single hydrogen bonds. SMD simulations further indicated that the R33A mutation weakens the hydrogen binding between all scaffold residues and MBP and not just between R33 and MBP. We validated the simulation data and characterized the effects of mutations on YS1-MBP binding by using single-molecule force spectroscopy and surface plasmon resonance. We propose that interfacial stability resulting from R33 of YS1 stacking with R344 of MBP synergistically stabilizes both its own bond and the interacting scaffold residues of YS1. Our integrated approach improves our understanding of the monobody scaffold interactions with a target, thus providing guidance for the improved engineering of monobodies. PMID:25650239

  19. Characterization of Monobody Scaffold Interactions with Ligand via Force Spectroscopy and Steered Molecular Dynamics

    NASA Astrophysics Data System (ADS)

    Cheung, Luthur Siu-Lun; Shea, Daniel J.; Nicholes, Nathan; Date, Amol; Ostermeier, Marc; Konstantopoulos, Konstantinos

    2015-02-01

    Monobodies are antibody alternatives derived from fibronectin that are thermodynamically stable, small in size, and can be produced in bacterial systems. Monobodies have been engineered to bind a wide variety of target proteins with high affinity and specificity. Using alanine-scanning mutagenesis simulations, we identified two scaffold residues that are critical to the binding interaction between the monobody YS1 and its ligand, maltose-binding protein (MBP). Steered molecular dynamics (SMD) simulations predicted that the E47A and R33A mutations in the YS1 scaffold substantially destabilize the YS1-MBP interface by reducing the bond rupture force and the lifetime of single hydrogen bonds. SMD simulations further indicated that the R33A mutation weakens the hydrogen binding between all scaffold residues and MBP and not just between R33 and MBP. We validated the simulation data and characterized the effects of mutations on YS1-MBP binding by using single-molecule force spectroscopy and surface plasmon resonance. We propose that interfacial stability resulting from R33 of YS1 stacking with R344 of MBP synergistically stabilizes both its own bond and the interacting scaffold residues of YS1. Our integrated approach improves our understanding of the monobody scaffold interactions with a target, thus providing guidance for the improved engineering of monobodies.

  20. Alternative Non-Antibody Protein Scaffolds for Molecular Imaging of Cancer

    PubMed Central

    Stern, Lawrence A.; Case, Brett A.; Hackel, Benjamin J.

    2013-01-01

    The development of improved methods for early detection and characterization of cancer presents a major clinical challenge. One approach that has shown excellent potential in preclinical and clinical evaluation is molecular imaging with small-scaffold, non-antibody based, engineered proteins. These novel diagnostic agents produce high contrast images due to their fast clearance from the bloodstream and healthy tissues, can be evolved to bind a multitude of cancer biomarkers, and are easily functionalized by site-specific bioconjugation methods. Several small protein scaffolds have been verified for in vivo molecular imaging including affibodies and their two-helix variants, knottins, fibronectins, DARPins, and several natural ligands. Further, the biodistribution of these engineered ligands can be optimized through rational mutation of the conserved regions, careful selection and placement of chelator, and modification of molecular size. PMID:24358455

  1. Experimental Tools to Study Molecular Recognition within the Nanoparticle Corona

    PubMed Central

    Landry, Markita P.; Kruss, Sebastian; Nelson, Justin T.; Bisker, Gili; Iverson, Nicole M.; Reuel, Nigel F.; Strano, Michael S.

    2014-01-01

    Advancements in optical nanosensor development have enabled the design of sensors using syntheticmolecular recognition elements through a recently developed method called Corona Phase MolecularRecognition (CoPhMoRe). The synthetic sensors resulting from these design principles are highly selective for specific analytes, and demonstrate remarkable stability for use under a variety of conditions. An essential element of nanosensor development hinges on the ability to understand the interface between nanoparticles and the associated corona phase surrounding the nanosensor, an environment outside of the range of traditional characterization tools, such as NMR. This review discusses the need for new strategies and instrumentation to study the nanoparticle corona, operating in both in vitro and in vivo environments. Approaches to instrumentation must have the capacity to concurrently monitor nanosensor operation and the molecular changes in the corona phase. A detailed overview of new tools for the understanding of CoPhMoRe mechanisms is provided for future applications. PMID:25184487

  2. Supramolecular recognition of estrogens via molecularly imprinted polymers

    PubMed Central

    Ričanyová, Júlia; Gadzała-Kopciuch, Renata; Szumski, Michał

    2010-01-01

    The isolation and preconcentration of estrogens from new types of biological samples (acellular and protein-free simulated body fluid) by molecularly imprinted solid-phase extraction has been described. In this technique, supramolecular receptors, namely molecularly imprinted polymers (MIPs) are used as a sorbent material. The recognition sites of MIPs were prepared by non-covalent multiple interactions and formed with the target 17β-estradiol as a template molecule. High-performance liquid chromatography with spectroscopic UV, selective, and a sensitive electrochemical CoulArray detector was used for the determination of 17β-estradiol, estrone, and estriol in simulated body fluid which mimicked human plasma. PMID:20549493

  3. Method of assembly of molecular-sized nets and scaffolding

    DOEpatents

    Michl, Josef; Magnera, Thomas F.; David, Donald E.; Harrison, Robin M.

    1999-01-01

    The present invention relates to methods and starting materials for forming molecular-sized grids or nets, or other structures based on such grids and nets, by creating molecular links between elementary molecular modules constrained to move in only two directions on an interface or surface by adhesion or bonding to that interface or surface. In the methods of this invention, monomers are employed as the building blocks of grids and more complex structures. Monomers are introduced onto and allowed to adhere or bond to an interface. The connector groups of adjacent adhered monomers are then polymerized with each other to form a regular grid in two dimensions above the interface. Modules that are not bound or adhered to the interface are removed prior to reaction of the connector groups to avoid undesired three-dimensional cross-linking and the formation of non-grid structures. Grids formed by the methods of this invention are useful in a variety of applications, including among others, for separations technology, as masks for forming regular surface structures (i.e., metal deposition) and as templates for three-dimensional molecular-sized structures.

  4. Method of assembly of molecular-sized nets and scaffolding

    DOEpatents

    Michl, J.; Magnera, T.F.; David, D.E.; Harrison, R.M.

    1999-03-02

    The present invention relates to methods and starting materials for forming molecular-sized grids or nets, or other structures based on such grids and nets, by creating molecular links between elementary molecular modules constrained to move in only two directions on an interface or surface by adhesion or bonding to that interface or surface. In the methods of this invention, monomers are employed as the building blocks of grids and more complex structures. Monomers are introduced onto and allowed to adhere or bond to an interface. The connector groups of adjacent adhered monomers are then polymerized with each other to form a regular grid in two dimensions above the interface. Modules that are not bound or adhered to the interface are removed prior to reaction of the connector groups to avoid undesired three-dimensional cross-linking and the formation of non-grid structures. Grids formed by the methods of this invention are useful in a variety of applications, including among others, for separations technology, as masks for forming regular surface structures (i.e., metal deposition) and as templates for three-dimensional molecular-sized structures. 9 figs.

  5. Orchestration of Molecular Information through Higher Order Chemical Recognition

    NASA Astrophysics Data System (ADS)

    Frezza, Brian M.

    Broadly defined, higher order chemical recognition is the process whereby discrete chemical building blocks capable of specifically binding to cognate moieties are covalently linked into oligomeric chains. These chains, or sequences, are then able to recognize and bind to their cognate sequences with a high degree of cooperativity. Principally speaking, DNA and RNA are the most readily obtained examples of this chemical phenomenon, and function via Watson-Crick cognate pairing: guanine pairs with cytosine and adenine with thymine (DNA) or uracil (RNA), in an anti-parallel manner. While the theoretical principles, techniques, and equations derived herein apply generally to any higher-order chemical recognition system, in practice we utilize DNA oligomers as a model-building material to experimentally investigate and validate our hypotheses. Historically, general purpose information processing has been a task limited to semiconductor electronics. Molecular computing on the other hand has been limited to ad hoc approaches designed to solve highly specific and unique computation problems, often involving components or techniques that cannot be applied generally in a manner suitable for precise and predictable engineering. Herein, we provide a fundamental framework for harnessing high-order recognition in a modular and programmable fashion to synthesize molecular information process networks of arbitrary construction and complexity. This document provides a solid foundation for routinely embedding computational capability into chemical and biological systems where semiconductor electronics are unsuitable for practical application.

  6. Magnetic molecularly imprinted polymer for aspirin recognition and controlled release

    NASA Astrophysics Data System (ADS)

    Kan, Xianwen; Geng, Zhirong; Zhao, Yao; Wang, Zhilin; Zhu, Jun-Jie

    2009-04-01

    Core-shell structural magnetic molecularly imprinted polymers (magnetic MIPs) with combined properties of molecular recognition and controlled release were prepared and characterized. Magnetic MIPs were synthesized by the co-polymerization of methacrylic acid (MAA) and trimethylolpropane trimethacrylate (TRIM) around aspirin (ASP) at the surface of double-bond-functionalized Fe3O4 nanoparticles in chloroform. The obtained spherical magnetic MIPs with diameters of about 500 nm had obvious superparamagnetism and could be separated quickly by an external magnetic field. Binding experiments were carried out to evaluate the properties of magnetic MIPs and magnetic non-molecularly imprinted polymers (magnetic NIPs). The results demonstrated that the magnetic MIPs had high adsorption capacity and selectivity to ASP. Moreover, release profiles and release rate of ASP from the ASP-loaded magnetic MIPs indicated that the magnetic MIPs also had potential applications in drug controlled release.

  7. The structural biology of molecular recognition by vancomycin.

    PubMed

    Loll, P J; Axelsen, P H

    2000-01-01

    Vancomycin is the archetype among naturally occurring compounds known as glycopeptide antibiotics. Because it is a vital therapeutic agent used world-wide for the treatment of infections with gram-positive bacteria, emerging bacterial resistance to vancomycin is a major public health threat. Recent investigations into the mechanisms of action of glycopeptide antibiotics are driven by a need to understand their detailed mechanism of action so that new agents can be developed to overcome resistance. These investigations have revealed that glycopeptide antibiotics exhibit a rich array of complex cooperative phenomena when they bind target ligands, making them valuable model systems for the study of molecular recognition. PMID:10940250

  8. Affinity sensor based on immobilized molecular imprinted synthetic recognition elements.

    PubMed

    Lenain, Pieterjan; De Saeger, Sarah; Mattiasson, Bo; Hedström, Martin

    2015-07-15

    An affinity sensor based on capacitive transduction was developed to detect a model compound, metergoline, in a continuous flow system. This system simulates the monitoring of low-molecular weight organic compounds in natural flowing waters, i.e. rivers and streams. During operation in such scenarios, control of the experimental parameters is not possible, which poses a true analytical challenge. A two-step approach was used to produce a sensor for metergoline. Submicron spherical molecularly imprinted polymers, used as recognition elements, were obtained through emulsion polymerization and subsequently coupled to the sensor surface by electropolymerization. This way, a robust and reusable sensor was obtained that regenerated spontaneously under the natural conditions in a river. Small organic compounds could be analyzed in water without manipulating the binding or regeneration conditions, thereby offering a viable tool for on-site application. PMID:25703726

  9. Fluorescence resonance energy transfer-based molecular logic circuit using a DNA scaffold

    NASA Astrophysics Data System (ADS)

    Nishimura, Takahiro; Ogura, Yusuke; Tanida, Jun

    2012-12-01

    This paper presents a method of information processing using biomolecular input signals and fluorescence resonance energy transfer (FRET) signaling constructed on a DNA scaffold. Logic operations are achieved by encoding molecular inputs into an arrangement of fluorescence dyes using simple DNA reactions and by evaluating a logic expression using local photonic signaling that is much faster than DNA reactions. Experimental results verify the operation of a complete set of Boolean logic functions (AND, OR, NOT) and combinational logic operations using a FRET-signal cascade.

  10. Quantum origins of molecular recognition and olfaction in drosophila

    NASA Astrophysics Data System (ADS)

    Bittner, Eric R.; Madalan, Adrian; Czader, Arkadiusz; Roman, Gregg

    2012-12-01

    The standard model for molecular recognition of an odorant is that receptor sites discriminate by molecular geometry as evidenced that two chiral molecules may smell very differently. However, recent studies of isotopically labeled olfactants indicate that there may be a molecular vibration-sensing component to olfactory reception, specifically in the spectral region around 2300 cm-1. Here, we present a donor-bridge-acceptor model for olfaction which attempts to explain this effect. Our model, based upon accurate quantum chemical calculations of the olfactant (bridge) in its neutral and ionized states, posits that internal modes of the olfactant are excited impulsively during hole transfer from a donor to acceptor site on the receptor, specifically those modes that are resonant with the tunneling gap. By projecting the impulsive force onto the internal modes, we can determine which modes are excited at a given value of the donor-acceptor tunneling gap. Only those modes resonant with the tunneling gap and are impulsively excited will give a significant contribution to the inelastic transfer rate. Using acetophenone as a test case, our model and experiments on D. melanogaster suggest that isotopomers of a given olfactant give rise to different odorant qualities. These results support the notion that inelastic scattering effects may play a role in discriminating between isotopomers but that this is not a general spectroscopic effect.

  11. Molecular Recognition of Gangliosides and Their Potential for Cancer Immunotherapies

    PubMed Central

    Krengel, Ute; Bousquet, Paula A.

    2014-01-01

    Gangliosides are sialic-acid-containing glycosphingolipids expressed on all vertebrate cells. They are primarily positioned in the plasma membrane with the ceramide part anchored in the membrane and the glycan part exposed on the surface of the cell. These lipids have highly diverse structures, not the least with respect to their carbohydrate chains, with N-acetylneuraminic acid (NeuAc) and N-glycolylneuraminic acid (NeuGc) being the two most common sialic-acid residues in mammalian cells. Generally, human healthy tissue is deficient in NeuGc, but this molecule is expressed in tumors and in human fetal tissues, and was hence classified as an onco-fetal antigen. Gangliosides perform important functions through carbohydrate-specific interactions with proteins, for example, as receptors in cell–cell recognition, which can be exploited by viruses and other pathogens, and also by regulating signaling proteins, such as the epidermal growth factor receptor (EGFR) and the vascular endothelial growth factor receptor (VEGFR), through lateral interaction in the membrane. Through both mechanisms, tumor-associated gangliosides may affect malignant progression, which makes them attractive targets for cancer immunotherapies. In this review, we describe how proteins recognize gangliosides, focusing on the molecular recognition of gangliosides associated with cancer immunotherapy, and discuss the importance of these molecules in cancer research. PMID:25101077

  12. Metabolite Recognition Principles and Molecular Mechanisms Underlying Riboswitch Function

    PubMed Central

    Serganov, Alexander; Patel, Dinshaw J.

    2015-01-01

    Riboswitches are mRNA elements capable of modulating gene expression in response to specific binding by cellular metabolites. Riboswitches exert their function through the interplay of alternative ligand-free and ligand-bound conformations of the metabolite-sensing domain, which in turn modulate the formation of adjacent gene expression controlling elements. X-ray crystallography and NMR spectroscopy have determined three-dimensional structures of virtually all the major riboswitch classes in the ligand-bound state and, for several riboswitches, in the ligand-free state. The resulting spatial topologies have demonstrated the wide diversity of riboswitch folds and revealed structural principles for specific recognition by cognate metabolites. The available three-dimensional information, supplemented by structure-guided biophysical and biochemical experimentation, has led to an improved understanding of how riboswitches fold, what RNA conformations are required for ligand recognition, and how ligand binding can be transduced into gene expression modulation. These studies have greatly facilitated the dissection of molecular mechanisms underlying riboswitch action and should in turn guide the anticipated development of tools for manipulating gene regulatory circuits. PMID:22577823

  13. Investigation of synthetic molecular recognition for biosensing applications

    NASA Astrophysics Data System (ADS)

    Stratis-Cullum, Dimitra N.; McMasters, Sun; Sooter, Letha J.; Pellegrino, Paul M.

    2007-04-01

    A fundamental understanding of the factors which influence binding performance is critical to any technology or methodology relying on molecular recognition of a specific target species. For the Army, there is a growing need for a basic understanding of these interactions with traditional recognition elements (e.g., antibodies) in non-traditional environmental conditions, such as with new and emerging threats. There is a similar need for building a base of knowledge on non-traditional affinity ligands that are biomimetic or biosynthetic in nature. In this paper, specific research at the Army Research Laboratory towards the development, evaluation and use of synthetic affinity ligands for sensing applications is discussed. This includes the results of our investigations of aptamer-based affinity ligands targeting Campylobacter jejuni. Using capillary electrophoretic techniques, the relative binding affinities of the aptamer ligands towards the target pathogen as well as the degree of cross-reactivity with other food borne-pathogens (i.e., Escherichia coli O157:H7 and Salmonella typhimurium) were evaluated. Current progress towards the development of synthetic affinity ligands for sensing applications will also be discussed.

  14. Protein flexibility and ligand recognition: challenges for molecular modeling.

    PubMed

    Spyrakis, Francesca; BidonChanal, Axel; Barril, Xavier; Luque, F Javier

    2011-01-01

    The intrinsic dynamics of macromolecules is an essential property to relate the structure of biomolecular systems with their function in the cell. In the field of ligand-receptor recognition, numerous evidences have revealed the limitations of the lock-and-key theory, and the need to elaborate models that take into account the inherent plasticity of biomolecules, such as the induced-fit model or the existence of an ensemble of pre-equilibrated conformations. Depending on the nature of the target system, ligand binding can be associated with small local adjustments in side chains or even the backbone to large-scale motions of structural fragments, domains or even subunits. Reproducing the inherent flexibility of biomolecules has thus become one of the most challenging issues in molecular modeling and simulation studies, as it has direct implications in our understanding of the structure-function relationships, but even in areas such as virtual screening and structure-based drug discovery. Given the intrinsic limitation of conventional simulation tools, only events occurring in short time scales can be reproduced at a high accuracy level through all-atom techniques such as Molecular Dynamics simulations. However, larger structural rearrangements demand the use of enhanced sampling methods relying on modified descriptions of the biomolecular system or the potential surface. This review illustrates the crucial role that structural plasticity plays in mediating ligand recognition through representative examples. In addition, it discusses some of the most powerful computational tools developed to characterize the conformational flexibility in ligand-receptor complexes. PMID:20939788

  15. Dynamics and functional differences between dendroaspin and rhodostomin: insights into protein scaffolds in integrin recognition.

    PubMed

    Cheng, Chun-Ho; Chen, Yi-Chun; Shiu, Jia-Hau; Chang, Yao-Tsung; Chang, Yung-Sheng; Huang, Chun-Hau; Chen, Chiu-Yueh; Chuang, Woei-Jer

    2012-12-01

    Dendroaspin (Den) and rhodostomin (Rho) are snake venom proteins containing a PRGDMP motif. Although Den and Rho have different 3D structures, they are highly potent integrin inhibitors. To study their structure, function, and dynamics relationships, we expressed Den and Rho in Pichia pastoris. The recombinant Den and Rho inhibited platelet aggregation with the K(I) values of 149.8 and 83.2 nM. Cell adhesion analysis showed that Den was 3.7 times less active than Rho when inhibiting the integrin αIIbβ3 and 2.5 times less active when inhibiting the integrin αvβ3. In contrast, Den and Rho were similarly active when inhibiting the integrin α5β1 with the IC₅₀ values of 239.8 and 256.8 nM. NMR analysis showed that recombinant Den and Rho have different 3D conformations for their arginyl-glycyl-aspartic acid (RGD) motif. However, the comparison with Rho showed that the docking of Den into integrin αvβ3 resulted in a similar number of contacts. Analysis of the dynamic properties of the RGD loop in Den and Rho showed that they also had different dynamic properties. These results demonstrate that protein scaffolds affect the function, structure, and dynamics of their RGD motif. PMID:23033223

  16. Dynamics and functional differences between dendroaspin and rhodostomin: Insights into protein scaffolds in integrin recognition

    PubMed Central

    Cheng, Chun-Ho; Chen, Yi-Chun; Shiu, Jia-Hau; Chang, Yao-Tsung; Chang, Yung-Sheng; Huang, Chun-Hau; Chen, Chiu-Yueh; Chuang, Woei-Jer

    2012-01-01

    Dendroaspin (Den) and rhodostomin (Rho) are snake venom proteins containing a PRGDMP motif. Although Den and Rho have different 3D structures, they are highly potent integrin inhibitors. To study their structure, function, and dynamics relationships, we expressed Den and Rho in Pichia pastoris. The recombinant Den and Rho inhibited platelet aggregation with the KI values of 149.8 and 83.2 nM. Cell adhesion analysis showed that Den was 3.7 times less active than Rho when inhibiting the integrin αIIbβ3 and 2.5 times less active when inhibiting the integrin αvβ3. In contrast, Den and Rho were similarly active when inhibiting the integrin α5β1 with the IC50 values of 239.8 and 256.8 nM. NMR analysis showed that recombinant Den and Rho have different 3D conformations for their arginyl-glycyl-aspartic acid (RGD) motif. However, the comparison with Rho showed that the docking of Den into integrin αvβ3 resulted in a similar number of contacts. Analysis of the dynamic properties of the RGD loop in Den and Rho showed that they also had different dynamic properties. These results demonstrate that protein scaffolds affect the function, structure, and dynamics of their RGD motif. PMID:23033223

  17. Molecular Recognition at the Protein-Hydroxyapatite Interface

    SciTech Connect

    Stayton, Partick S.; Drobny, G. P.; Shaw, Wendy J.; Long, Joanna R.; Gilbert, Michelle R.

    2003-09-01

    Proteins found in mineralized tissues act as nature's crystal engineers, where they play a key role in promoting or inhibiting the growth of minerals, such as hydroxyapitite (bones/teeth) and calcium oxalate (kidney stones). Despite their importance in hard-tissue formation and remodeling, and in pathological processes such as stone formation and arterial calcification, there is little known of the protein structure-function relationships that govern hard-tissue engineering. Here we review early studies that have utilized solid-state NMR (ssNMR) techniques to provide in situ secondary-structure determination of statherin and statherin peptides on their biologically relevant hydroxyapatite (HAP) surfaces. In addition to direct structural study, molecular dynamics studies have provided considerable insight into the protein-binding footprint on hydroxyapatite. The molecular insight provided by these studies has also led to the design of biomimetic fusion peptides that utilize nature's crystal-recognition mechanism to display accessible and dynamic bioactive sequences from the HAP surface. These peptides selectively engage adhesion receptors and direct specific outside-in signaling pathway activation in osteoblast-like cells.

  18. A divergent synthetic approach to diverse molecular scaffolds: assessment of lead-likeness using LLAMA, an open-access computational tool.

    PubMed

    Colomer, Ignacio; Empson, Christopher J; Craven, Philip; Owen, Zachary; Doveston, Richard G; Churcher, Ian; Marsden, Stephen P; Nelson, Adam

    2016-06-01

    Complementary cyclisation reactions of hex-2-ene-1,6-diamine derivatives were exploited in the synthesis of alternative molecular scaffolds. The value of the synthetic approach was analysed using LLAMA, an open-access computational tool for assessing the lead-likeness and novelty of molecular scaffolds. PMID:27145833

  19. Comparison of two molecular scaffolds, 5-methylisoxazole-3-carboxamide and 5-methylisoxazole-4-carboxamide.

    PubMed

    Song, Yaoming; Zhang, Yiguan; Lee, An-Rong; Huang, Wen-Hsin; Chen, Ben; Palfey, Bruce; Shaw, Jiajiu

    2014-01-01

    Leflunomide is a disease-modifying antirheumatic drug (DMARD) for the treatment of rheumatoid arthritis (RA). Structurally, it is a derivative of 5-methylisoxazole-4-carboxamide. Upon metabolism, the N-O bond in the isoxazole ring is cleaved to form the active metabolite, teriflunomide, which was recently approved by the FDA for the treatment of multiple sclerosis. Both leflunomide and teriflunomide inhibit dihydroorotate dehydrogenase (DHODH) thereby inhibiingt the synthesis of pyrimidine. For both drugs, the two major concerns are potential liver toxicity and teratogenicity. It was suspected that these undesirable effects might be related to the cleavage of the N-O bond. We herein summarize the metabolites-toxicity issues related to leflunomide/teriflunomide and discuss two related molecular platforms, UTL-4 and UTL-5. UTL-4 compounds are based on the same scaffold of leflunomide; their toxicological and pharmacological effects are not significantly different from those of leflunomide/teriflunomide. In UTL-5 series, the leflunomide scaffold is changed into 5-methylisoxazole-3-carboxamide. Unlike leflunomide, the N-O bond of a UTL-5 compound, UTL-5b, is not cleaved upon metabolism; instead, the peptide bond is cleaved to form its major metabolites. UTL-5b and its metabolites do not inhibit DHODH in vitro. In addition, UTL-5b and all other UTL-5 compounds have lower acute toxicity than leflunomide/teriflunomide. Furthermore, from leflunomide to UTL-5b/UTL-5g, the potential liver toxicity becomes liver protective effect. With the reduced toxicity, UTL-5 compounds still maintain significant pharmacological effects including anti-inflammatory and antiarthritic effects. In summary, our observations provide a valuable direction in drug optimization based on the modification of the leflunomide scaffold. PMID:23944378

  20. Comparison of multi-recognition molecularly imprinted polymers for recognition of melamine, cyromazine, triamterene, and trimethoprim.

    PubMed

    Wang, Xian-Hua; Zhang, Jing; Peng, Chao; Dong, Qian; Huang, Yan-Ping; Liu, Zhao-Sheng

    2015-09-01

    Three fragmental templates, including 2,4-diamino-6-methyl-1,3,5-triazine (DMT), cyromazine (CYR), and trimethoprim (TME), were used to prepare the fragment molecularly imprinted polymers (FMIPs), respectively, in polar ternary porogen which was composed of ionic liquid ([BMIM]BF4), methanol, and water. The morphology, specific surface areas, and selectivity of the obtained FMIPs for fragmental analogues were systematically characterized. The experimental results showed that the FMIPs possessed the best specific recognition ability to the relative template and the greatest imprinting factor (IF) was 5.25, 6.69, and 7.11 of DMT on DMT-MIPs, CYR on CYR-MIPs, and TME on TME-MIPs, respectively. In addition, DMT-MIPs also showed excellent recognition capability for fragmental analogues including CYR, melamine (MEL), triamterene (TAT), and TME, and the IFs were 2.08, 3.89, 2.18, and 2.60, respectively. The effects of pH and temperature on the retention of the fragmental and structural analogues were studied in detail. Van't Hoff analysis indicated that the retention and selectivity on FMIPs were an entropy-driven process, i.e., steric interaction. The resulting DMT-MIPs were used as a solid-phase extraction material to enrich CYR, MEL, TAT, and TME in different bio-matrix samples for high-performance liquid chromatography analysis. The developed method had acceptable recoveries (86.8-98.6%, n = 3) and precision (2.7-4.6%) at three spiked levels (0.05-0.5 μg g(-1)). PMID:26195027

  1. Nanoparticle-Templated Molecular Recognition Platforms for Detection of Biological Analytes.

    PubMed

    Beyene, Abraham G; Demirer, Gozde S; Landry, Markita P

    2016-01-01

    Molecular recognition of biological analytes with optical nanosensors provides both spatial and temporal biochemical information. A recently developed sensing platform exploits near-infrared fluorescent single-wall carbon nanotubes combined with electrostatically pinned heteropolymers to yield a synthetic molecular recognition technique that is maximally transparent through biological matter. This molecular recognition technique is known as corona phase molecular recognition (CoPhMoRe). In CoPhMoRe, the specificity of a folded polymer toward an analyte does not arise from a pre-existing polymer-analyte chemical affinity. Rather, specificity is conferred through conformational changes undergone by a polymer that is pinned to the surface of a nanoparticle in the presence of an analyte and the subsequent modifications in fluorescence readout of the nanoparticles. The protocols in this article describe a novel single-molecule microscopy tool (near-infrared fluorescence and total internal reflection fluorescence [nIRF TIRF] hybrid microscope) to visualize the CoPhMoRe recognition process, enabling a better understanding of synthetic molecular recognition. We describe this requisite microscope for simultaneous single-molecule visualization of optical molecular recognition and signal transduction. We elaborate on the general procedures for synthesizing and identifying single-walled carbon nanotube-based sensors that employ CoPhMoRe via two biologically relevant examples of single-molecule recognition for the hormone estradiol and the neurotransmitter dopamine. © 2016 by John Wiley & Sons, Inc. PMID:27622569

  2. The DBHS proteins SFPQ, NONO and PSPC1: a multipurpose molecular scaffold.

    PubMed

    Knott, Gavin J; Bond, Charles S; Fox, Archa H

    2016-05-19

    Nuclear proteins are often given a concise title that captures their function, such as 'transcription factor,' 'polymerase' or 'nuclear-receptor.' However, for members of the Drosophila behavior/human splicing (DBHS) protein family, no such clean-cut title exists. DBHS proteins are frequently identified engaging in almost every step of gene regulation, including but not limited to, transcriptional regulation, RNA processing and transport, and DNA repair. Herein, we present a coherent picture of DBHS proteins, integrating recent structural insights on dimerization, nucleic acid binding modalities and oligomerization propensity with biological function. The emerging paradigm describes a family of dynamic proteins mediating a wide range of protein-protein and protein-nucleic acid interactions, on the whole acting as a multipurpose molecular scaffold. Overall, significant steps toward appreciating the role of DBHS proteins have been made, but we are only beginning to understand the complexity and broader importance of this family in cellular biology. PMID:27084935

  3. Molecular Recognition of Platinated DNA from Chromosomal HMGB1.

    PubMed

    Nguyen, Trung Hai; Rossetti, Giulia; Arnesano, Fabio; Ippoliti, Emiliano; Natile, Giovanni; Carloni, Paolo

    2014-08-12

    Cisplatin cures testicular and ovarian cancers with unprecedented potency. It induces its beneficial activity by covalently binding to DNA. Repair enzymes, which remove the platinated lesions from DNA, cause drug resistance. Chromosomal High Mobility Group Box proteins (HMGB) may interfere with this process by binding to platinated DNA. Using 8 μs multiple-walker well-tempered metadynamics simulations, here, we investigated the structural and the energetic determinants of one of the HMGB proteins (HMGB1A) in complex with the platinated oligonucleotide [Pt(NH3)2](2+)-d(CCUCTCTG*G*ACCTTCC)-d(GGAGAGACCTGGAAGG) (*G are platinated guanines), for which experimental structural information is available. The calculated affinity is in good agreement with experiment. The process is predicted to be enthalpy-driven, as found for other protein/DNA complexes. The Lys7 residue, whose side-chain was not resolved in the X-ray structure, is found to interact with the C4 5'-phosphate and this interaction emerges as a key facet for the molecular recognition process. In addition, our calculations provide a molecular basis for the experimentally measured decreased affinity of HMGB1A for platinated DNA, as a consequence of Cys22-Cys44 S-S bridge formation (such an oxidation cannot take place in some members of this protein family present in the testis, where the drug is particularly effective). This decrease is likely to be caused by a small yet significant rearrangement of helices H1 and H2 with consequent alteration of the Phe37 juxtaposition. PMID:26588321

  4. Molecularly imprinted silica-silver nanowires for tryptophan recognition

    NASA Astrophysics Data System (ADS)

    Díaz-Faes López, T.; Díaz-García, M. E.; Badía-Laíño, R.

    2014-10-01

    We report on silver nanowires (AgNWs) coated with molecularly imprinted silica (MIP SiO2) for recognition of tryptophan (Trp). The use of AgNWs as a template confers an imprinted material with adequate mechanical strength and with a capability of recognizing Trp due to its nanomorphology when compared to spherical microparticles with a similar surface-to-volume ratio. Studies on adsorption isotherms showed the MIP-SiO2-AgNWs to exhibit homogeneous affinity sites with narrow affinity distribution. This suggests that the synthesized material behaves as a 1D nanomaterial with a large area and small thickness with very similar affinity sites. Trp release from MIP-SiO2-AgNWs was demonstrated to be dominated by the diffusion rate of Trp as controlled by the specific interactions with the imprinted silica shell. Considering these results and the lack of toxicity of silica sol-gel materials, the material offers potential in the field of drug or pharmaceutical controlled delivery, but also in optoelectronic devices, electrodes and sensors.

  5. Flexibility and molecular recognition in the immune system

    PubMed Central

    Jimenez, Ralph; Salazar, Georgina; Baldridge, Kim K.; Romesberg, Floyd E.

    2003-01-01

    Photon echo spectroscopy has been used to measure the response of three antibody-binding sites to perturbation from electronic excitation of a bound antigen, fluorescein. The three antibodies show motions that range in time scale from tens of femtoseconds to nanoseconds. Relative to the others, one antibody, 4-4-20, possesses a rigid binding site that likely results from a short and inflexible heavy chain complementarity-determining region 3 (HCDR3) loop and a critical Tyr that acts as a “molecular splint,” rigidifying the antigen across its most flexible internal degree of freedom. The remaining two antibodies, 34F10 and 40G4, despite being generated against the same antigen, possess binding sites that are considerably more flexible. The more flexible combining sites likely result from longer HCDR3 loops and a deletion in the light chain complementarity-determining region 1 (LCDR1) that removes the critical Tyr residue. The binding site flexibilities may result in varying mechanisms of antigen recognition including lock-and-key, induced-fit, and conformational selection. PMID:12518056

  6. Aptamers: versatile molecular recognition probes for cancer detection

    PubMed Central

    Sun, Hongguang; Tan, Weihong; Zu, Youli

    2015-01-01

    In the past two decades, aptamers have emerged as a novel class of molecular recognition probes comprising uniquely-folded short RNA or single-stranded DNA oligonucleotides that bind to their cognate targets with high specificity and affinity. Aptamers, often referred to as “chemical antibodies”, possess several highly desirable features for clinical use. They can be chemically synthesized and are easily conjugated to a wide range of reporters for different applications, and are able to rapidly penetrate tissues. These advantages significantly enhance their clinical applicability, and render them excellent alternatives to antibody-based probes in cancer diagnostics and therapeutics. Aptamer probes based on fluorescence, colorimetry, magnetism, electrochemistry, and in conjunction with nanomaterials (e.g., nanoparticles, quantum dots, single-walled carbon nanotubes, and magnetic nanoparticles) have provided novel ultrasensitive cancer diagnostic strategies and assays. Furthermore, promising aptamer targeted-multimodal tumor imaging probes have been recently developed in conjunction with fluorescence, positron emission tomography (PET), single-photon emission computed tomography (SPECT), and magnetic resonance imaging (MRI). The capabilities of the aptamer-based platforms described herein underscore the great potential they hold for the future of cancer detection. In this review, we highlight the most prominent recent developments in this rapidly advancing field. PMID:26618445

  7. Use of ultra-high molecular weight polycaprolactone scaffolds for ACL reconstruction.

    PubMed

    Leong, Natalie L; Kabir, Nima; Arshi, Armin; Nazemi, Azadeh; Jiang, Jie; Wu, Ben M; Petrigliano, Frank A; McAllister, David R

    2016-05-01

    Previously, we reported on the implantation of electrospun polycaprolactone (PCL) grafts for use in ACL tissue engineering in a small animal model. In the present study, we hypothesized that grafts fabricated from ultra-high molecular weight polycaprolactone (UHMWPCL) would have similarly favorable biologic properties but superior mechanical properties as compared to grafts fabricated from PCL. Two forms of polycaprolactone were obtained (UHMWPCL, MW = 500 kD, and PCL, MW = 80 kD) and electrospun into scaffolds that were used to perform ACL reconstruction in 7-8 week old male Lewis rats. The following groups were examined: UHMWPCL, PCL, flexor digitorum longus (FDL) allograft, native ACL, as well as sham surgery in which the ACL was transsected. At 16 weeks post-operatively, biomechanical testing, histology, and immunohistochemistry (IHC) were performed. Analysis of cellularity indicated that there was no significant difference among the UHMWPCL, PCL, and FDL allograft groups. Quantification of birefringence from picrosirius red staining demonstrated significantly more aligned collagen fibers in the allograft than the PCL group, but no difference between the UHMWPCL and allograft groups. The peak load to failure of the UHMWPCL grafts was significantly higher than PCL, and not significantly different from FDL allograft. This in vivo study establishes the superiority of the higher molecular weight version of polycaprolactone over PCL as a scaffold material for ACL reconstruction. By 16 weeks after implantation, the UHMWPCL grafts were not significantly different from the FDL allografts in terms of cellularity, peak load to failure, stiffness, and collagen fiber alignment. © 2015 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 34:828-835, 2016. PMID:26497133

  8. Dissecting the molecular origins of specific protein-nucleic acid recognition: hydrostatic pressure and molecular dynamics.

    PubMed Central

    Lynch, Thomas W; Kosztin, Dorina; McLean, Mark A; Schulten, Klaus; Sligar, Stephen G

    2002-01-01

    The fundamental processes by which proteins recognize and bind to nucleic acids are critical to understanding cellular function. To explore the factors involved in protein-DNA recognition, we used hydrostatic pressure to perturb the binding of the BamHI endonuclease to cognate DNA, both in experiment and in molecular dynamic simulations. A new technique of high-pressure gel mobility shift analysis was used to test the effects of elevated hydrostatic pressure on the binding of BamHI to its cognate recognition sequence. Upon application of a pressure of 500 bar, the equilibrium dissociation constant of BamHI binding to the cognate site was found to increase nearly 10-fold. A challenge has been to link this type of pure thermodynamic measurement to functional events occurring at the molecular level. Thus, we used molecular dynamic simulations at both ambient and elevated pressures to reveal details of the direct and water-mediated interactions between BamHI and cognate DNA, which allow explanation of the effects of pressure on site-specific protein-DNA binding and complex stability. PMID:11751298

  9. Novel thiol-based histone deacetylase inhibitors bearing 3-phenyl-1H-pyrazole-5-carboxamide scaffold as surface recognition motif: Design, synthesis and SAR study.

    PubMed

    Wen, Jiachen; Niu, Qun; Liu, Jiang; Bao, Yu; Yang, Jinyu; Luan, Shenglin; Fan, Yinbo; Liu, Dan; Zhao, Linxiang

    2016-01-15

    A series of novel thiol-based histone deacetylase (HDAC) inhibitors bearing 3-phenyl-1H-pyrazole-5-carboxamide scaffold as surface recognition motif was designed, synthesized, and evaluated for their HDAC inhibition activity. Among them, 15j (IC50=0.08μM) was identified as a better inhibitor than Vorinostat (IC50=0.25μM) against total HDACs. In addition, Structure-activity relationships (SAR) analyses indicated that (i) compounds with different substituents on pyrazole N-1 position exhibited superior activities than those on pyrazole N-2 position, (ii) variation of functional groups on N-1'-alkyl chain terminus followed the trends of carboxyl group>hydroxyl group≫alkyl group, and (iii) methylation on pyrazole C-4 position diminished the HDAC inhibition activity. The SAR will guide us to further refine compounds bearing 3-phenyl-1H-pyrazole-5-carboxamide scaffold to achieve better HDAC inhibitors. PMID:26706171

  10. Phage randomization in a charybdotoxin scaffold leads to CD4-mimetic recognition motifs that bind HIV-1 envelope through non-aromatic sequences.

    PubMed

    Li, C; Dowd, C S; Zhang, W; Chaiken, I M

    2001-06-01

    Binding of HIV-1 gp120 to T-cell receptor CD4 initiates conformational changes in the viral envelope that trigger viral entry into host cells. Phage epitope randomization of a beta-turn loop of a charybdotoxin-based miniprotein scaffold was used to identify peptides that can bind gp120 and block the gp120-CD4 interaction. We describe here the display of the charybdotoxin scaffold on the filamentous phage fUSE5, its use to construct a beta-turn library, and miniprotein sequences identified through library panning with immobilized Env gp120. Competition enzyme-linked immunosorbent assay (ELISA) identified high-frequency phage selectants for which specific gp120 binding was competed by sCD4. Several of these selectants contain hydrophobic residues in place of the Phe that occurs in the gp120-binding beta-turns of both CD4 and previously identified scorpion toxin CD4 mimetics. One of these selectants, denoted TXM[24GQTL27], contains GQTL in place of the CD4 beta-turn sequence 40QGSF43. TXM[24GQTL27] peptide was prepared using solid-phase chemical synthesis, its binding to gp120 demonstrated by optical biosensor kinetics analysis and its affinity for the CD4 binding site of gp120 confirmed by competition ELISA. The results demonstrate that aromatic-less loop-containing CD4 recognition mimetics can be formed with detectable envelope protein binding within a beta-turn of the charybdotoxin miniprotein scaffold. The results of this work establish a methodology for phage display of a charybdotoxin miniprotein scaffold and point to the potential value of phage-based epitope randomization of this miniprotein for identifying novel CD4 mimetics. The latter are potentially useful in deconvoluting structural determinants of CD4-HIV envelope recognition and possibly in designing antagonists of viral entry. PMID:11437954

  11. Molecular tips for scanning tunneling microscopy: intermolecular electron tunneling for single-molecule recognition and electronics.

    PubMed

    Nishino, Tomoaki

    2014-01-01

    This paper reviews the development of molecular tips for scanning tunneling microscopy (STM). Molecular tips offer many advantages: first is their ability to perform chemically selective imaging because of chemical interactions between the sample and the molecular tip, thus improving a major drawback of conventional STM. Rational design of the molecular tip allows sophisticated chemical recognition; e.g., chiral recognition and selective visualization of atomic defects in carbon nanotubes. Another advantage is that they provide a unique method to quantify electron transfer between single molecules. Understanding such electron transfer is mandatory for the realization of molecular electronics. PMID:24420248

  12. Carbon Nanomaterials and DNA: from Molecular Recognition to Applications.

    PubMed

    Sun, Hanjun; Ren, Jinsong; Qu, Xiaogang

    2016-03-15

    DNA is polymorphic. Increasing evidence has indicated that many biologically important processes are related to DNA's conformational transition and assembly states. In particular, noncanonical DNA structures, such as the right-handed A-form, the left-handed Z-form, the triplex, the G-quadruplex, the i-motif, and so forth, have been specific targets for the diagnosis and therapy of human diseases. Meanwhile, they have been widely used in the construction of smart DNA nanomaterials and nanoarchitectures. As rising stars in materials science, the family of carbon nanomaterials (CNMs), including two-dimensional graphene, one-dimensional carbon nanotubes (CNTs), and zero-dimensional graphene or carbon quantum dots (GQDs or CQDs), interact with DNA and are able to regulate the conformational transitions of DNA. The interaction of DNA with CNMs not only opens new opportunities for specific molecular recognition, but it also expands the promising applications of CNMs from materials science to biotechnology and biomedicine. In this Account, we focus on our contributions to the field of interactions between CNMs and DNA in which we have explored their promising applications in nanodevices, sensing, materials synthesis, and biomedicine. For one-dimensional CNTs, two-dimensional graphene, and zero-dimensional GQDs and CQDs, the basic principles, binding modes, and applications of the interactions between CNMs and DNA are reviewed. We aim to give prominence to the important status of CNMs in the field of molecular recognition for DNA. First, we summarized our discovery of the interactions between single-walled carbon nanotubes (SWNTs) with duplex, triplex, and human telomeric i-motif DNA and their interesting applications. For example, SWNTs are the first chemical agents that can selectively stabilize human telomeric i-motif DNA and induce its formation under physiological conditions. On the basis of this principle, two types of nanodevices were designed. One was used for

  13. Molecularly Imprinted Polymers: Thermodynamic and Kinetic Considerations on the Specific Sorption and Molecular Recognition

    PubMed Central

    Li, Songjun; Huang, Xing; Zheng, Mingxia; Li, Wuke; Tong, Kejun

    2008-01-01

    This article presents a work aiming at thermodynamically and kinetically interpreting the specific sorption and recognition by a molecularly imprinted polymer. Using Boc-L-Phe-OH as a template, the imprinted material was prepared. The result indicates that the prepared polymer can well discriminate the imprint species from its analogue (Boc-D-Phe-OH), so as to adsorb more for the former but less for the latter. Kinetic analysis indicates that this specific sorption, in nature, can be a result of a preferential promotion. The imprint within the polymer causes a larger adsorption rate for the template than for the analogue. Thermodynamic study also implies that the molecular induction from the specific imprint to the template is larger than to the analogue, which thus makes the polymer capable of preferentially alluring the template to bind.

  14. ION AND MOLECULE SENSORS USING MOLECULAR RECOGNITION IN LUMINESCENT, CONDUCTIVE POLYMERS

    EPA Science Inventory

    This program integrates three individual, highly interactive projects that will use molecular recognition strategies to develop sensor technology based on luminescent, conductive polymers that contain sites for binding specific molecules or ions in the presence of related molecul...

  15. Molecular Recognition of Biomolecules by Chiral CdSe Quantum Dots.

    PubMed

    Mukhina, Maria V; Korsakov, Ivan V; Maslov, Vladimir G; Purcell-Milton, Finn; Govan, Joseph; Baranov, Alexander V; Fedorov, Anatoly V; Gun'ko, Yurii K

    2016-01-01

    Molecular recognition is one of the most important phenomena in Chemistry and Biology. Here we present a new way of enantiomeric molecular recognition using intrinsically chiral semiconductor nanocrystals as assays. Real-time confocal microscopy studies supported by circular dichroism spectroscopy data and theoretical modelling indicate an ability of left-handed molecules of cysteine and, to a smaller extent, histidine and arginine to discriminate between surfaces of left- and right-handed nanocrystals. PMID:27063962

  16. Molecular Recognition of Biomolecules by Chiral CdSe Quantum Dots

    NASA Astrophysics Data System (ADS)

    Mukhina, Maria V.; Korsakov, Ivan V.; Maslov, Vladimir G.; Purcell-Milton, Finn; Govan, Joseph; Baranov, Alexander V.; Fedorov, Anatoly V.; Gun’Ko, Yurii K.

    2016-04-01

    Molecular recognition is one of the most important phenomena in Chemistry and Biology. Here we present a new way of enantiomeric molecular recognition using intrinsically chiral semiconductor nanocrystals as assays. Real-time confocal microscopy studies supported by circular dichroism spectroscopy data and theoretical modelling indicate an ability of left-handed molecules of cysteine and, to a smaller extent, histidine and arginine to discriminate between surfaces of left- and right-handed nanocrystals.

  17. Molecular Recognition of Biomolecules by Chiral CdSe Quantum Dots

    PubMed Central

    Mukhina, Maria V.; Korsakov, Ivan V.; Maslov, Vladimir G.; Purcell-Milton, Finn; Govan, Joseph; Baranov, Alexander V.; Fedorov, Anatoly V.; Gun’ko, Yurii K.

    2016-01-01

    Molecular recognition is one of the most important phenomena in Chemistry and Biology. Here we present a new way of enantiomeric molecular recognition using intrinsically chiral semiconductor nanocrystals as assays. Real-time confocal microscopy studies supported by circular dichroism spectroscopy data and theoretical modelling indicate an ability of left-handed molecules of cysteine and, to a smaller extent, histidine and arginine to discriminate between surfaces of left- and right-handed nanocrystals. PMID:27063962

  18. The Integration of 3-D Cell-Printing and Mesoscopic Fluorescence Molecular Tomography of Vascular Constructs within Thick Hydrogel Scaffolds

    PubMed Central

    Zhao, Lingling; Lee, Vivian K.; Yoo, Seung-Schik; Dai, Guohao; Intes, Xavier

    2012-01-01

    Developing methods that provide adequate vascular perfusion is an important step toward engineering large functional tissues. Meanwhile, an imaging modality to assess the three-dimensional (3-D) structures and functions of the vascular channels is lacking for thick matrices (>2~3mm). Herein, we report on an original approach to construct and image 3-D dynamically perfused vascular structures in thick hydrogel scaffolds. In this work, we integrated a robotic 3-D cell-printing technology with a mesoscopic fluorescence molecular tomography imaging system, and demonstrated the capability of the platform to construct perfused collagen scaffolds with endothelial lining and to image both the fluid flow and fluorescent-labeled living endothelial cells at high-frame rates, with high sensitivity and accuracy. These results establish the potential of integrating both 3-D cell-printing and fluorescence mesoscopic imaging for functional and molecular studies in complex tissue engineered tissues. PMID:22531221

  19. The DBHS proteins SFPQ, NONO and PSPC1: a multipurpose molecular scaffold

    PubMed Central

    Knott, Gavin J.; Bond, Charles S.; Fox, Archa H.

    2016-01-01

    Nuclear proteins are often given a concise title that captures their function, such as ‘transcription factor,’ ‘polymerase’ or ‘nuclear-receptor.’ However, for members of the Drosophila behavior/human splicing (DBHS) protein family, no such clean-cut title exists. DBHS proteins are frequently identified engaging in almost every step of gene regulation, including but not limited to, transcriptional regulation, RNA processing and transport, and DNA repair. Herein, we present a coherent picture of DBHS proteins, integrating recent structural insights on dimerization, nucleic acid binding modalities and oligomerization propensity with biological function. The emerging paradigm describes a family of dynamic proteins mediating a wide range of protein–protein and protein–nucleic acid interactions, on the whole acting as a multipurpose molecular scaffold. Overall, significant steps toward appreciating the role of DBHS proteins have been made, but we are only beginning to understand the complexity and broader importance of this family in cellular biology. PMID:27084935

  20. Syntheses of steroid-based molecularly imprinted polymers and their molecular recognition study with spectrometric detection

    NASA Astrophysics Data System (ADS)

    Dong, He; Tong, Ai-jun; Li, Long-di

    2003-01-01

    Recognition of five steroid compounds, β-estradiol, ethynylestradiol, estradiolbenzoate, testosterone and methyltestosterone were studied using a synthesized molecularly imprinted polymer (MIP). When β-estradiol was used as the template molecule, the polymer was synthesized with methacrylic acid (MAA) as the functional monomer and ethylene glycol dimethacrylate (EGDMA) as the cross linking agent through non-covalent interactions. It is found that the kind of porogen solvent and the polymerization conditions greatly affected the binding ability of a MIP to a certain molecule. Releasing of the template was performed by continuous extraction with methanol containing 10% acetic acid in a Soxhlet extractor. Our results indicated that such carefully synthesized MIP showed specific affinity toward β-estradiol in the adsorption process.

  1. Corona Phase Molecular Recognition (CoPhMoRe) to Enable New Nanosensor Interfaces

    NASA Astrophysics Data System (ADS)

    Strano, Michael

    2015-03-01

    Our lab at MIT has been interested in how the 1D and 2D electronic structures of carbon nanotubes and graphene respectively can be utilized to advance new concepts in molecular detection. We introduce CoPhMoRe or corona phase molecular recognition as a method of discovering synthetic antibodies, or nanotube-templated recognition sites from a heteropolymer library. We show that certain synthetic heteropolymers, once constrained onto a single-walled carbon nanotube by chemical adsorption, also form a new corona phase that exhibits highly selective recognition for specific molecules. To prove the generality of this phenomenon, we report three examples of heteropolymers-nanotube recognition complexes for riboflavin, L-thyroxine and estradiol. The platform opens new opportunities to create synthetic recognition sites for molecular detection. We have also extended this molecular recognition technique to neurotransmitters, producing the first fluorescent sensor for dopamine. Another area of advancement in biosensor development is the use of near infrared fluorescent carbon nanotube sensors for in-vivo detection. Here, we show that PEG-ligated d(AAAT)7 DNA wrapped SWNT are selective for nitric oxide, a vasodilator of blood vessels, and can be tail vein injected into mice and localized within the viable mouse liver. We use an SJL mouse model to study liver inflammation in vivo using the spatially and spectrally resolved nIR signature of the localized SWNT sensors.

  2. Protected Amine Labels: A Versatile Molecular Scaffold for Multiplexed Nominal Mass and Sub-Da Isotopologue Quantitative Proteomic Reagents

    NASA Astrophysics Data System (ADS)

    Ficarro, Scott B.; Biagi, Jessica M.; Wang, Jinhua; Scotcher, Jenna; Koleva, Rositsa I.; Card, Joseph D.; Adelmant, Guillaume; He, Huan; Askenazi, Manor; Marshall, Alan G.; Young, Nicolas L.; Gray, Nathanael S.; Marto, Jarrod A.

    2014-04-01

    We assemble a versatile molecular scaffold from simple building blocks to create binary and multiplexed stable isotope reagents for quantitative mass spectrometry. Termed Protected Amine Labels (PAL), these reagents offer multiple analytical figures of merit including, (1) robust targeting of peptide N-termini and lysyl side chains, (2) optimal mass spectrometry ionization efficiency through regeneration of primary amines on labeled peptides, (3) an amino acid-based mass tag that incorporates heavy isotopes of carbon, nitrogen, and oxygen to ensure matched physicochemical and MS/MS fragmentation behavior among labeled peptides, and (4) a molecularly efficient architecture, in which the majority of hetero-atom centers can be used to synthesize a variety of nominal mass and sub-Da isotopologue stable isotope reagents. We demonstrate the performance of these reagents in well-established strategies whereby up to four channels of peptide isotopomers, each separated by 4 Da, are quantified in MS-level scans with accuracies comparable to current commercial reagents. In addition, we utilize the PAL scaffold to create isotopologue reagents in which labeled peptide analogs differ in mass based on the binding energy in carbon and nitrogen nuclei, thereby allowing quantification based on MS or MS/MS spectra. We demonstrate accurate quantification for reagents that support 6-plex labeling and propose extension of this scheme to 9-channels based on a similar PAL scaffold. Finally, we provide exemplar data that extend the application of isotopologe-based quantification reagents to medium resolution, quadrupole time-of-flight mass spectrometers.

  3. Molecular Recognition of Insulin by a Synthetic Receptor

    SciTech Connect

    Chinai, Jordan M.; Taylor, Alexander B.; Ryno, Lisa M.; Hargreaves, Nicholas D.; Morris, Christopher A.; Hart, P. John; Urbach, Adam R.

    2011-08-29

    The discovery of molecules that bind tightly and selectively to desired proteins continues to drive innovation at the interface of chemistry and biology. This paper describes the binding of human insulin by the synthetic receptor cucurbit[7]uril (Q7) in vitro. Isothermal titration calorimetry and fluorescence spectroscopy experiments show that Q7 binds to insulin with an equilibrium association constant of 1.5 x 10{sup 6} M{sup -1} and with 50-100-fold selectivity versus proteins that are much larger but lack an N-terminal aromatic residue, and with >1000-fold selectivity versus an insulin variant lacking the N-terminal phenylalanine (Phe) residue. The crystal structure of the Q7{center_dot}insulin complex shows that binding occurs at the N-terminal Phe residue and that the N-terminus unfolds to enable binding. These findings suggest that site-selective recognition is based on the properties inherent to a protein terminus, including the unique chemical epitope presented by the terminal residue and the greater freedom of the terminus to unfold, like the end of a ball of string, to accommodate binding. Insulin recognition was predicted accurately from studies on short peptides and exemplifies an approach to protein recognition by targeting the terminus.

  4. Molecular basis for convergent evolution of glutamate recognition by pentameric ligand-gated ion channels

    PubMed Central

    Lynagh, Timothy; Beech, Robin N.; Lalande, Maryline J.; Keller, Kevin; Cromer, Brett A.; Wolstenholme, Adrian J.; Laube, Bodo

    2015-01-01

    Glutamate is an indispensable neurotransmitter, triggering postsynaptic signals upon recognition by postsynaptic receptors. We questioned the phylogenetic position and the molecular details of when and where glutamate recognition arose in the glutamate-gated chloride channels. Experiments revealed that glutamate recognition requires an arginine residue in the base of the binding site, which originated at least three distinct times according to phylogenetic analysis. Most remarkably, the arginine emerged on the principal face of the binding site in the Lophotrochozoan lineage, but 65 amino acids upstream, on the complementary face, in the Ecdysozoan lineage. This combined experimental and computational approach throws new light on the evolution of synaptic signalling. PMID:25708000

  5. Molecular recognition of human ephrinB2 cell surface receptor by an emergent African henipavirus

    PubMed Central

    Lee, Benhur; Pernet, Olivier; Ahmed, Asim A.; Zeltina, Antra; Beaty, Shannon M.; Bowden, Thomas A.

    2015-01-01

    The discovery of African henipaviruses (HNVs) related to pathogenic Hendra virus (HeV) and Nipah virus (NiV) from Southeast Asia and Australia presents an open-ended health risk. Cell receptor use by emerging African HNVs at the stage of host-cell entry is a key parameter when considering the potential for spillover and infection of human populations. The attachment glycoprotein from a Ghanaian bat isolate (GhV-G) exhibits <30% sequence identity with Asiatic NiV-G/HeV-G. Here, through functional and structural analysis of GhV-G, we show how this African HNV targets the same human cell-surface receptor (ephrinB2) as the Asiatic HNVs. We first characterized this virus−receptor interaction crystallographically. Compared with extant HNV-G–ephrinB2 structures, there was significant structural variation in the six-bladed β-propeller scaffold of the GhV-G receptor-binding domain, but not the Greek key fold of the bound ephrinB2. Analysis revealed a surprisingly conserved mode of ephrinB2 interaction that reflects an ongoing evolutionary constraint among geographically distal and phylogenetically divergent HNVs to maintain the functionality of ephrinB2 recognition during virus–host entry. Interestingly, unlike NiV-G/HeV-G, we could not detect binding of GhV-G to ephrinB3. Comparative structure–function analysis further revealed several distinguishing features of HNV-G function: a secondary ephrinB2 interaction site that contributes to more efficient ephrinB2-mediated entry in NiV-G relative to GhV-G and cognate residues at the very C terminus of GhV-G (absent in Asiatic HNV-Gs) that are vital for efficient receptor-induced fusion, but not receptor binding per se. These data provide molecular-level details for evaluating the likelihood of African HNVs to spill over into human populations. PMID:25825759

  6. Molecular recognition of human ephrinB2 cell surface receptor by an emergent African henipavirus.

    PubMed

    Lee, Benhur; Pernet, Olivier; Ahmed, Asim A; Zeltina, Antra; Beaty, Shannon M; Bowden, Thomas A

    2015-04-28

    The discovery of African henipaviruses (HNVs) related to pathogenic Hendra virus (HeV) and Nipah virus (NiV) from Southeast Asia and Australia presents an open-ended health risk. Cell receptor use by emerging African HNVs at the stage of host-cell entry is a key parameter when considering the potential for spillover and infection of human populations. The attachment glycoprotein from a Ghanaian bat isolate (GhV-G) exhibits <30% sequence identity with Asiatic NiV-G/HeV-G. Here, through functional and structural analysis of GhV-G, we show how this African HNV targets the same human cell-surface receptor (ephrinB2) as the Asiatic HNVs. We first characterized this virus-receptor interaction crystallographically. Compared with extant HNV-G-ephrinB2 structures, there was significant structural variation in the six-bladed β-propeller scaffold of the GhV-G receptor-binding domain, but not the Greek key fold of the bound ephrinB2. Analysis revealed a surprisingly conserved mode of ephrinB2 interaction that reflects an ongoing evolutionary constraint among geographically distal and phylogenetically divergent HNVs to maintain the functionality of ephrinB2 recognition during virus-host entry. Interestingly, unlike NiV-G/HeV-G, we could not detect binding of GhV-G to ephrinB3. Comparative structure-function analysis further revealed several distinguishing features of HNV-G function: a secondary ephrinB2 interaction site that contributes to more efficient ephrinB2-mediated entry in NiV-G relative to GhV-G and cognate residues at the very C terminus of GhV-G (absent in Asiatic HNV-Gs) that are vital for efficient receptor-induced fusion, but not receptor binding per se. These data provide molecular-level details for evaluating the likelihood of African HNVs to spill over into human populations. PMID:25825759

  7. Molecular recognition of organic ammonium ions in solution using synthetic receptors

    PubMed Central

    Späth, Andreas

    2010-01-01

    Summary Ammonium ions are ubiquitous in chemistry and molecular biology. Considerable efforts have been undertaken to develop synthetic receptors for their selective molecular recognition. The type of host compounds for organic ammonium ion binding span a wide range from crown ethers to calixarenes to metal complexes. Typical intermolecular interactions are hydrogen bonds, electrostatic and cation–π interactions, hydrophobic interactions or reversible covalent bond formation. In this review we discuss the different classes of synthetic receptors for organic ammonium ion recognition and illustrate the scope and limitations of each class with selected examples from the recent literature. The molecular recognition of ammonium ions in amino acids is included and the enantioselective binding of chiral ammonium ions by synthetic receptors is also covered. In our conclusion we compare the strengths and weaknesses of the different types of ammonium ion receptors which may help to select the best approach for specific applications. PMID:20502608

  8. Transfer of molecular recognition information from DNA nanostructures to gold nanoparticles

    NASA Astrophysics Data System (ADS)

    Edwardson, Thomas G. W.; Lau, Kai Lin; Bousmail, Danny; Serpell, Christopher J.; Sleiman, Hanadi F.

    2016-02-01

    DNA nanotechnology offers unparalleled precision and programmability for the bottom-up organization of materials. This approach relies on pre-assembling a DNA scaffold, typically containing hundreds of different strands, and using it to position functional components. A particularly attractive strategy is to employ DNA nanostructures not as permanent scaffolds, but as transient, reusable templates to transfer essential information to other materials. To our knowledge, this approach, akin to top-down lithography, has not been examined. Here we report a molecular printing strategy that chemically transfers a discrete pattern of DNA strands from a three-dimensional DNA structure to a gold nanoparticle. We show that the particles inherit the DNA sequence configuration encoded in the parent template with high fidelity. This provides control over the number of DNA strands and their relative placement, directionality and sequence asymmetry. Importantly, the nanoparticles produced exhibit the site-specific addressability of DNA nanostructures, and are promising components for energy, information and biomedical applications.

  9. The Molecular Basis of N-End Rule Recognition

    SciTech Connect

    Wang, K.H.; Roman-Hernandez, G.; Grant, R.A.; Sauer, R.T.; Baker, T.A.

    2009-03-27

    The N-end rule targets specific proteins for destruction in prokaryotes and eukaryotes. Here, we report a crystal structure of a bacterial N-end rule adaptor, ClpS, bound to a peptide mimic of an N-end rule substrate. This structure, which was solved at a resolution of 1.15 {angstrom}, reveals specific recognition of the peptide {alpha}-amino group via hydrogen bonding and shows that the peptide's N-terminal tyrosine side chain is buried in a deep hydrophobic cleft that pre-exists on the surface of ClpS. The adaptor side chains that contact the peptide's N-terminal residue are highly conserved in orthologs and in E3 ubiquitin ligases that mediate eukaryotic N-end rule recognition. We show that mutation of critical ClpS contact residues abrogates substrate delivery to and degradation by the AAA+ protease ClpAP, demonstrate that modification of the hydrophobic pocket results in altered N-end rule specificity, and discuss functional implications for the mechanism of substrate delivery.

  10. The Molecular Basis of N-end Rule Recognition

    PubMed Central

    Wang, Kevin H.; Roman-Hernandez, Giselle; Grant, Robert A.; Sauer, Robert T.; Baker, Tania A.

    2011-01-01

    Summary The N-end rule targets specific proteins for destruction in prokaryotes and eukaryotes. Here, we report a crystal structure of a bacterial N-end rule adaptor, ClpS, bound to a peptide mimic of an N-end rule substrate. This structure, which was solved at a resolution of 1.15 Å, reveals specific recognition of the peptide α-amino group via hydrogen bonding and shows that the peptide’s N-terminal tyrosine side chain is buried in a deep hydrophobic cleft that preexists on the surface of ClpS. The adaptor side chains that contact the peptide’s N-terminal residue are highly conserved in orthologs and in E3 ubiquitin ligases that mediate eukaryotic N-end rule recognition. We show that mutation of critical ClpS contact residues abrogates substrate delivery to and degradation by the AAA+ protease ClpAP, demonstrate that modification of the hydrophobic pocket results in altered N-end rule specificity, and discuss functional implications for the mechanism of substrate delivery. PMID:18995838

  11. Multipoint molecular recognition within a calix[6]arene funnel complex

    PubMed Central

    Coquière, David; de la Lande, Aurélien; Martí, Sergio; Parisel, Olivier; Prangé, Thierry; Reinaud, Olivia

    2009-01-01

    A multipoint recognition system based on a calix[6]arene is described. The calixarene core is decorated on alternating aromatic subunits by 3 imidazole arms at the small rim and 3 aniline groups at the large rim. This substitution pattern projects the aniline nitrogens toward each other when Zn(II) binds at the Tris-imidazole site or when a proton binds at an aniline. The XRD structure of the monoprotonated complex having an acetonitrile molecule bound to Zn(II) in the cavity revealed a constrained geometry at the metal center reminiscent of an entatic state. Computer modeling suggests that the aniline groups behave as a tritopic monobasic site in which only 1 aniline unit is protonated and interacts with the other 2 through strong hydrogen bonding. The metal complex selectively binds a monoprotonated diamine vs. a monoamine through multipoint recognition: coordination to the metal ion at the small rim, hydrogen bonding to the calix-oxygen core, CH/π interaction within the cavity's aromatic walls, and H-bonding to the anilines at the large rim. PMID:19237564

  12. Strategy for molecular beacon binding readout: separating molecular recognition element and signal reporter.

    PubMed

    Wang, Yongxiang; Li, Jishan; Jin, Jianyu; Wang, Hao; Tang, Hongxing; Yang, Ronghua; Wang, Kemin

    2009-12-01

    A new strategy for molecular beacon binding readout is proposed by using separation of the molecular recognition element and signal reporter. The signal transduction of the target binding event is based on displacing interaction between the target DNA and a competitor, the signal transducer. The target-free capture DNA is first interacted with the competitor, forming an assembled complex. In the presence of a target DNA that the affinity is stronger than that of the competitor, hybridization between capture DNA and the target disassembles the assembled complex and releases the free competitor to change the readout of the signal reporter. To demonstrate the feasibility of the design, a thymine-rich oligonucleotide was examined as a model system. Hg2+ was selected as the competitor, and mercaptoacetic acid-coated CdTe/ZnS quantum dots served as the fluorescent reporter. Selective binding of Hg2+ between the two thymine bases of the capture DNA forms a hairpin-structure. Hybridization between the capture DNA and target DNA destroys the hairpin-structure, releasing Hg2+ ions to quench the quantum dots fluorescence. Under the optimal conditions, fluorescence intensity of the quantum dots against the concentration of perfect cDNA was linear over the concentration range of 0.1-1.6 microM, with a limit of detection of 25 nM. This new assay method is simple in design, avoiding any oligonucleotide labeling. Furthermore, this strategy is generalizable since any target binding can in principle release the signal transducer and be detected with separated signal reporter. PMID:19899746

  13. Effects of substitution groups of glutamide-derived molecular gels on molecular shape recognition.

    PubMed

    Noguchi, Hiroki; Charoenraks, Tiraporn; Takafuji, Makoto; Ihara, Hirotaka

    2015-05-01

    We have reported that self-assembling glutamide lipid-grafted porous silica particles show high selectivity towards polycyclic aromatic hydrocarbons (PAHs) and bio-molecules. This enhancement of molecular recognition is brought about by the formation of a highly ordered structure in the glutamide lipid through intermolecular hydrogen bonding. To utilise, for selective separations, the highly oriented structure of glutamide lipids on the silica surface, in this study, we synthesised four glutamide lipids with different substitution groups and studied the separation behaviours of the glutamide lipid-grafted porous silica particles as reversed phase high-performance liquid chromatography (RP-HPLC) stationary phases. According to the HPLC studies, the functional group substitutions and the spacer group connecting the glutamide moiety and silica surface had a noticeable effect on the retention behaviours. Particularly, glutamide lipids with a long spacer group (C10) showed a higher selectivity towards positional (o-/m-/p-terphenyl) and geometrical (cis-/trans-stilbene) isomers. Differential scanning calorimetric and Fourier transform infrared spectroscopic studies suggested that the long spacer group induced the assembled structure of the glutamide lipid on the silica surface. Interestingly, the glutamide lipids with dodecyl groups and benzyl groups showed reverse elution orders towards the length to breadth ratio of PAHs. PMID:25817707

  14. Molecular Recognition and Structural Influences on Function in Bio-nanosystems of Nucleic Acids and Proteins

    NASA Astrophysics Data System (ADS)

    Sethaphong, Latsavongsakda

    This work examines smart material properties of rational self-assembly and molecular recognition found in nano-biosystems. Exploiting the sequence and structural information encoded within nucleic acids and proteins will permit programmed synthesis of nanomaterials and help create molecular machines that may carry out new roles involving chemical catalysis and bioenergy. Responsive to different ionic environments thru self-reorgnization, nucleic acids (NA) are nature's signature smart material; organisms such as viruses and bacteria use features of NAs to react to their environment and orchestrate their lifecycle. Furthermore, nucleic acid systems (both RNA and DNA) are currently exploited as scaffolds; recent applications have been showcased to build bioelectronics and biotemplated nanostructures via directed assembly of multidimensional nanoelectronic devices 1. Since the most stable and rudimentary structure of nucleic acids is the helical duplex, these were modeled in order to examine the influence of the microenvironment, sequence, and cation-dependent perturbations of their canonical forms. Due to their negatively charged phosphate backbone, NA's rely on counterions to overcome the inherent repulsive forces that arise from the assembly of two complementary strands. As a realistic model system, we chose the HIV-TAR helix (PDB ID: 397D) to study specific sequence motifs on cation sequestration. At physiologically relevant concentrations of sodium and potassium ions, we observed sequence based effects where purine stretches were adept in retaining high residency cations. The transitional space between adenine and guanosine nucleotides (ApG step) in a sequence proved the most favorable. This work was the first to directly show these subtle interactions of sequence based cationic sequestration and may be useful for controlling metallization of nucleic acids in conductive nanowires. Extending the study further, we explored the degree to which the structure of NA

  15. How much do van der Waals dispersion forces contribute to molecular recognition in solution?

    NASA Astrophysics Data System (ADS)

    Yang, Lixu; Adam, Catherine; Nichol, Gary S.; Cockroft, Scott L.

    2013-12-01

    The emergent properties that arise from self-assembly and molecular recognition phenomena are a direct consequence of non-covalent interactions. Gas-phase measurements and computational methods point to the dominance of dispersion forces in molecular association, but solvent effects complicate the unambiguous quantification of these forces in solution. Here, we have used synthetic molecular balances to measure interactions between apolar alkyl chains in 31 organic, fluorous and aqueous solvent environments. The experimental interaction energies are an order of magnitude smaller than estimates of dispersion forces between alkyl chains that have been derived from vaporization enthalpies and dispersion-corrected calculations. Instead, it was found that cohesive solvent-solvent interactions are the major driving force behind apolar association in solution. The results suggest that theoretical models that implicate important roles for dispersion forces in molecular recognition events should be interpreted with caution in solvent-accessible systems.

  16. Small molecule recognition of mephedrone using an anthracene molecular clip.

    PubMed

    Kellett, Kathryn; Broome, J Hugh; Zloh, Mire; Kirton, Stewart B; Fergus, Suzanne; Gerhard, Ute; Stair, Jacqueline L; Wallace, Karl J

    2016-06-14

    An anthracene molecular probe has been synthesised and shown to target mephedrone, a stimulant drug from the cathinone class of new psychoactive substances (NPS). A protocol has been developed to detect mephedrone via the probe using NMR spectroscopy in a simulated street sample containing two of the most common cutting agents, benzocaine and caffeine. PMID:27198990

  17. Molecular recognition and self-assembly special feature: Multipoint molecular recognition within a calix[6]arene funnel complex.

    PubMed

    Coquière, David; de la Lande, Aurélien; Martí, Sergio; Parisel, Olivier; Prangé, Thierry; Reinaud, Olivia

    2009-06-30

    A multipoint recognition system based on a calix[6]arene is described. The calixarene core is decorated on alternating aromatic subunits by 3 imidazole arms at the small rim and 3 aniline groups at the large rim. This substitution pattern projects the aniline nitrogens toward each other when Zn(II) binds at the Tris-imidazole site or when a proton binds at an aniline. The XRD structure of the monoprotonated complex having an acetonitrile molecule bound to Zn(II) in the cavity revealed a constrained geometry at the metal center reminiscent of an entatic state. Computer modeling suggests that the aniline groups behave as a tritopic monobasic site in which only 1 aniline unit is protonated and interacts with the other 2 through strong hydrogen bonding. The metal complex selectively binds a monoprotonated diamine vs. a monoamine through multipoint recognition: coordination to the metal ion at the small rim, hydrogen bonding to the calix-oxygen core, CH/pi interaction within the cavity's aromatic walls, and H-bonding to the anilines at the large rim. PMID:19237564

  18. Molecular Recognition of Agonist and Antagonist for Peroxisome Proliferator-Activated Receptor-α Studied by Molecular Dynamics Simulations

    PubMed Central

    Liu, Mengyuan; Wang, Lushan; Zhao, Xian; Sun, Xun

    2014-01-01

    Peroxisome proliferator activated receptor-α (PPAR-α) is a ligand-activated transcription factor which plays important roles in lipid and glucose metabolism. The aim of this work is to find residues which selectively recognize PPAR-α agonists and antagonists. To achieve this aim, PPAR-α/13M and PPAR-α/471 complexes were subjected to perform molecular dynamics simulations. This research suggests that several key residues only participate in agonist recognition, while some other key residues only contribute to antagonist recognition. It is hoped that such work is useful for medicinal chemists to design novel PPAR-α agonists and antagonists. PMID:24837836

  19. LARGE SCALE EVALUATION OF A PATTERN RECOGNITION/EXPERT SYSTEM FOR MASS SPECTRAL MOLECULAR WEIGHT ESTIMATION

    EPA Science Inventory

    A fast, personal-computer based method of estimating molecular weights of organic compounds from low resolution mass I spectra has been thoroughly evaluated. he method is based on a rule-based pattern,recognition/expert system approach which uses empirical linear corrections whic...

  20. Molecular recognition of nitrated fatty acids by PPAR[gamma

    SciTech Connect

    Li, Yong; Zhang, Jifeng; Schopfer, Francisco J.; Martynowski, Dariusz; Garcia-Barrio, Minerva T.; Kovach, Amanda; Suino-Powell, Kelly; Baker, Paul R.S.; Freeman, Bruce A.; Chen, Y. Eugene; Xu, H. Eric

    2010-03-08

    Peroxisome proliferator activated receptor-{gamma} (PPAR{gamma}) regulates metabolic homeostasis and adipocyte differentiation, and it is activated by oxidized and nitrated fatty acids. Here we report the crystal structure of the PPAR{gamma} ligand binding domain bound to nitrated linoleic acid, a potent endogenous ligand of PPAR{gamma}. Structural and functional studies of receptor-ligand interactions reveal the molecular basis of PPAR{gamma} discrimination of various naturally occurring fatty acid derivatives.

  1. Molecular mechanism underlying promiscuous polyamine recognition by spermidine acetyltransferase.

    PubMed

    Sugiyama, Shigeru; Ishikawa, Sae; Tomitori, Hideyuki; Niiyama, Mayumi; Hirose, Mika; Miyazaki, Yuma; Higashi, Kyohei; Murata, Michio; Adachi, Hiroaki; Takano, Kazufumi; Murakami, Satoshi; Inoue, Tsuyoshi; Mori, Yusuke; Kashiwagi, Keiko; Igarashi, Kazuei; Matsumura, Hiroyoshi

    2016-07-01

    Spermidine acetyltransferase (SAT) from Escherichia coli, which catalyses the transfer of acetyl groups from acetyl-CoA to spermidine, is a key enzyme in controlling polyamine levels in prokaryotic cells. In this study, we determined the crystal structure of SAT in complex with spermidine (SPD) and CoA at 2.5Å resolution. SAT is a dodecamer organized as a hexamer of dimers. The secondary structural element and folding topology of the SAT dimer resemble those of spermidine/spermine N(1)-acetyltransferase (SSAT), suggesting an evolutionary link between SAT and SSAT. However, the polyamine specificity of SAT is distinct from that of SSAT and is promiscuous. The SPD molecule is also located at the inter-dimer interface. The distance between SPD and CoA molecules is 13Å. A deep, highly acidic, water-filled cavity encompasses the SPD and CoA binding sites. Structure-based mutagenesis and in-vitro assays identified SPD-bound residues, and the acidic residues lining the walls of the cavity are mostly essential for enzymatic activities. Based on mutagenesis and structural data, we propose an acetylation mechanism underlying promiscuous polyamine recognition for SAT. PMID:27163532

  2. Systematic assessment of scaffold hopping versus activity cliff formation across bioactive compound classes following a molecular hierarchy.

    PubMed

    Stumpfe, Dagmar; Dimova, Dilyana; Bajorath, Jürgen

    2015-07-01

    Scaffold hopping and activity cliff formation define opposite ends of the activity landscape feature spectrum. To rationalize these events at the level of scaffolds, active compounds involved in scaffold hopping were required to contain topologically distinct scaffolds but have only limited differences in potency, whereas compounds involved in activity cliffs were required to share the same scaffold but have large differences in potency. A systematic search was carried out for compounds involved in scaffold hopping and/or activity cliff formation. Results obtained for compound data sets covering more than 300 human targets revealed clear trends. If scaffolds represented multiple but fewer than 10 active compounds, nearly 90% of all scaffolds were exclusively involved in hopping events. With increasing compound coverage, the fraction of scaffolds involved in both scaffold hopping and activity cliff formation significantly increased to more than 50%. However, ∼40% of the scaffolds representing large numbers of active compounds continued to be exclusively involved in scaffold hopping. More than 200 scaffolds with broad target coverage were identified that consistently represented potent compounds and yielded an abundance of scaffold hops in the low-nanomolar range. These and other subsets of scaffolds we characterized are of prime interest for structure-activity relationship (SAR) exploration and compound design. Therefore, the complete scaffold classification generated in the course of our analysis is made freely available. PMID:25982076

  3. Protected Amine Labels: A Versatile Molecular Scaffold for Multiplexed Nominal Mass and Sub-Da Isotopologue Quantitative Proteomic Reagents

    PubMed Central

    Ficarro, Scott B.; Biagi, Jessica M.; Wang, Jinhua; Scotcher, Jenna; Koleva, Rositsa I.; Card, Joseph D.; Adelmant, Guillaume; He, Huan; Askenazi, Manor; Marshall, Alan G.; Young, Nicolas L.; Gray, Nathanael S.; Marto, Jarrod A.

    2014-01-01

    We assemble a versatile molecular scaffold from simple building blocks to create binary and multiplexed stable isotope reagents for quantitative mass spectrometry. Termed Protected Amine Labels (PAL), these reagents offer multiple analytical figures of merit including, (i) robust targeting of peptide N-termini and lysyl side chains, (ii) optimal mass spectrometry ionization efficiency through regeneration of primary amines on labeled peptides, (iii) an amino acid-based mass tag that incorporates heavy isotopes of carbon, nitrogen, and oxygen to ensure matched physicochemical and MS/MS fragmentation behavior among labeled peptides, and (iv) a molecularly efficient architecture, in which the majority of hetero-atom centers can be used to synthesize a variety of nominal mass and sub-Da isotopologue stable isotope reagents. We demonstrate the performance of these reagents in well-established strategies whereby up to four channels of peptide isotopomers, each separated by 4 Da are quantified in MS-level scans with accuracies comparable to current commercial reagents. In addition we utilize the PAL scaffold to create isotopologue reagents in which labeled peptide analogs differ in mass based on the binding energy in carbon and nitrogen nuclei, thereby allowing quantification based on MS or MS/MS spectra. We demonstrate accurate quantification for reagents that support 6-plex labeling and propose extension of this scheme to 9-channels based on a similar PAL scaffold. Finally we provide exemplar data that extends the application of isotopologe-based quantification reagents to medium resolution, quadrupole time-of-flight mass spectrometers. PMID:24496597

  4. Protected amine labels: a versatile molecular scaffold for multiplexed nominal mass and sub-Da isotopologue quantitative proteomic reagents.

    PubMed

    Ficarro, Scott B; Biagi, Jessica M; Wang, Jinhua; Scotcher, Jenna; Koleva, Rositsa I; Card, Joseph D; Adelmant, Guillaume; He, Huan; Askenazi, Manor; Marshall, Alan G; Young, Nicolas L; Gray, Nathanael S; Marto, Jarrod A

    2014-04-01

    We assemble a versatile molecular scaffold from simple building blocks to create binary and multiplexed stable isotope reagents for quantitative mass spectrometry. Termed Protected Amine Labels (PAL), these reagents offer multiple analytical figures of merit including, (1) robust targeting of peptide N-termini and lysyl side chains, (2) optimal mass spectrometry ionization efficiency through regeneration of primary amines on labeled peptides, (3) an amino acid-based mass tag that incorporates heavy isotopes of carbon, nitrogen, and oxygen to ensure matched physicochemical and MS/MS fragmentation behavior among labeled peptides, and (4) a molecularly efficient architecture, in which the majority of hetero-atom centers can be used to synthesize a variety of nominal mass and sub-Da isotopologue stable isotope reagents. We demonstrate the performance of these reagents in well-established strategies whereby up to four channels of peptide isotopomers, each separated by 4 Da, are quantified in MS-level scans with accuracies comparable to current commercial reagents. In addition, we utilize the PAL scaffold to create isotopologue reagents in which labeled peptide analogs differ in mass based on the binding energy in carbon and nitrogen nuclei, thereby allowing quantification based on MS or MS/MS spectra. We demonstrate accurate quantification for reagents that support 6-plex labeling and propose extension of this scheme to 9-channels based on a similar PAL scaffold. Finally, we provide exemplar data that extend the application of isotopologe-based quantification reagents to medium resolution, quadrupole time-of-flight mass spectrometers. PMID:24496597

  5. Molecular recognition of α-cyclodextrin (CD) to choral amino acids based on methyl orange as a molecular probe

    NASA Astrophysics Data System (ADS)

    Yuexian, Fan; Yu, Yang; Shaomin, Shuang; Chuan, Dong

    2005-03-01

    The molecular recognition interaction of α-CD to chiral amino acids was investigated by using spectrophotometry based on methyl orange as a molecular probe. The molecular recognition ability depended on the inclusion formation constants. The molecular recognition of α-CD to aromatic amino acids was the order: DL-tryptophan > L-tryptophan > L-phenylalanine > L-tyrosine ≈ DL-β-3,4-dihydroxy-phenylalanine; whereas for aliphatic amino acids, the order was: L- iso-leucine > L-leucine ≈ L-methionine ≈ DL-mehtionine > D-leucine. The effect of temperature on the inclusion interaction was examined and the thermodynamic parameters of inclusion process, Δ G, Δ H, Δ S, were determined. The experimental results indicated that the inclusion process was an exothermic and enthalpy-driven process accompanied with a negative or minor positive entropic contribution. The inclusion interaction between α-CD and amino acids satisfied the law of enthalpy-entropy compensation. The compensation temperature was 291 K.

  6. Molecular recognition of alpha-cyclodextrin (CD) to choral amino acids based on methyl orange as a molecular probe.

    PubMed

    Yuexian, Fan; Yu, Yang; Shaomin, Shuang; Chuan, Dong

    2005-03-01

    The molecular recognition interaction of alpha-CD to chiral amino acids was investigated by using spectrophotometry based on methyl orange as a molecular probe. The molecular recognition ability depended on the inclusion formation constants. The molecular recognition of alpha-CD to aromatic amino acids was the order: DL-tryptophan > L-tryptophan > L-phenylalanine > L-tyrosine approximately DL-beta-3,4-dihydroxy-phenylalanine; whereas for aliphatic amino acids, the order was: L-iso-leucine > L-leucine approximately L-methionine approximately DL-mehtionine > D-leucine. The effect of temperature on the inclusion interaction was examined and the thermodynamic parameters of inclusion process, delta G, delta H, delta S, were determined. The experimental results indicated that the inclusion process was an exothermic and enthalpy-driven process accompanied with a negative or minor positive entropic contribution. The inclusion interaction between alpha-CD and amino acids satisfied the law of enthalpy-entropy compensation. The compensation temperature was 291 K. PMID:15683802

  7. ERK Signals: Scaffolding Scaffolds?

    PubMed Central

    Casar, Berta; Crespo, Piero

    2016-01-01

    ERK1/2 MAP Kinases become activated in response to multiple intra- and extra-cellular stimuli through a signaling module composed of sequential tiers of cytoplasmic kinases. Scaffold proteins regulate ERK signals by connecting the different components of the module into a multi-enzymatic complex by which signal amplitude and duration are fine-tuned, and also provide signal fidelity by isolating this complex from external interferences. In addition, scaffold proteins play a central role as spatial regulators of ERKs signals. In this respect, depending on the subcellular localization from which the activating signals emanate, defined scaffolds specify which substrates are amenable to be phosphorylated. Recent evidence has unveiled direct interactions among different scaffold protein species. These scaffold-scaffold macro-complexes could constitute an additional level of regulation for ERK signals and may serve as nodes for the integration of incoming signals and the subsequent diversification of the outgoing signals with respect to substrate engagement. PMID:27303664

  8. Autophagic clearance of bacterial pathogens: molecular recognition of intracellular microorganisms

    PubMed Central

    Mansilla Pareja, Maria Eugenia; Colombo, Maria I.

    2013-01-01

    Autophagy is involved in several physiological and pathological processes. One of the key roles of the autophagic pathway is to participate in the first line of defense against the invasion of pathogens, as part of the innate immune response. Targeting of intracellular bacteria by the autophagic machinery, either in the cytoplasm or within vacuolar compartments, helps to control bacterial proliferation in the host cell, controlling also the spreading of the infection. In this review we will describe the means used by diverse bacterial pathogens to survive intracellularly and how they are recognized by the autophagic molecular machinery, as well as the mechanisms used to avoid autophagic clearance. PMID:24137567

  9. How hydrophobic drying forces impact the kinetics of molecular recognition.

    PubMed

    Mondal, Jagannath; Morrone, Joseph A; Berne, B J

    2013-08-13

    A model of protein-ligand binding kinetics, in which slow solvent dynamics results from hydrophobic drying transitions, is investigated. Molecular dynamics simulations show that solvent in the receptor pocket can fluctuate between wet and dry states with lifetimes in each state that are long enough for the extraction of a separable potential of mean force and wet-to-dry transitions. We present a diffusive surface hopping model that is represented by a 2D Markovian master equation. One dimension is the standard reaction coordinate, the ligand-pocket separation, and the other is the solvent state in the region between ligand and binding pocket which specifies whether it is wet or dry. In our model, the ligand diffuses on a dynamic free-energy surface which undergoes kinetic transitions between the wet and dry states. The model yields good agreement with results from explicit solvent molecular dynamics simulation and an improved description of the kinetics of hydrophobic assembly. Furthermore, it is consistent with a "non-Markovian Brownian theory" for the ligand-pocket separation coordinate alone. PMID:23901110

  10. Energetics, Thermodynamics, and Molecular Recognition of Piperine with DNA.

    PubMed

    Haris, P; Mary, Varughese; Haridas, M; Sudarsanakumar, C

    2015-12-28

    Piperine, the bioactive phytochemical from black pepper (Piper nigrum L.), is a nontoxic natural compound exhibiting many physiological and pharmacological properties. They include antioxidant, anti-inflammatory, antimutagenic, antitumor, antiapoptotic, antigenotoxic, antiarthritic, antifungal, antimicrobial, antidepressant, anti-HBV, and gastro-protective activities. It also enhances the bioavailability of phytochemicals and drugs. The molecular mechanism of action of piperine with DNA has not yet been addressed, while its pharmacological activities have been reported. In this work we report for the first time the interaction of piperine molecule with DNA duplex. We have carried out UV-vis absorption and fluorescence spectroscopy to confirm the binding of piperine with calf thymus DNA (ctDNA). The energetics of interaction of piperine with ctDNA was monitored by isothermal titration calorimetry (ITC). Differential scanning calorimetry (DSC) and melting temperature (Tm) analysis were also performed, confirming a minor groove mode of binding of piperine with ctDNA. The binding free energy ΔG values obtained from molecular dynamics simulation studies agree well with ITC values and reveal a sequence dependent minor groove binding exhibiting a specificity toward AT rich sequences. PMID:26523930

  11. Adsorption and recognition characteristics of surface molecularly imprinted polymethacrylic acid/silica toward genistein.

    PubMed

    Zhang, Yanyan; Gao, Baojiao; An, Fuqiang; Xu, Zeqing; Zhang, Tingting

    2014-09-12

    In this paper, on the basis of surface-initiated graft polymerization, a new surface molecular imprinting technique is established by molecular design. And molecularly imprinted polymer MIP-PMAA/SiO2 is successfully prepared with genistein as template. The adsorption and recognition characteristics of MIP-PMAA/SiO2 for genistein are studied in depth by using static method, dynamic method and competitive adsorption experiment. The experimental results show that MIP-PMAA/SiO2 possesses very strong adsorption affinity and specific recognition for genistein. The saturated adsorption capacity could reach to 0.36mmolg(-1). The selectivity coefficients relative to quercetin and rutin are 5.4 and 11.8, respectively. Besides, MIP-PMAA/SiO2 is regenerated easily and exhibits excellent reusability. PMID:25085816

  12. Design, synthesis and decoration of molecular scaffolds for exploitation in the production of alkaloid-like libraries.

    PubMed

    Craven, Philip; Aimon, Anthony; Dow, Mark; Fleury-Bregeot, Nicolas; Guilleux, Rachel; Morgentin, Remy; Roche, Didier; Kalliokoski, Tuomo; Foster, Richard; Marsden, Stephen P; Nelson, Adam

    2015-06-01

    The design, synthesis and decoration of six small molecule libraries is described. Each library was inspired by structures embedded in the framework of specific alkaloid natural products. The development of optimised syntheses of the required molecular scaffolds is described, in which reactions including Pd-catalysed aminoarylation and diplolar cycloadditions have been exploited as key steps. The synthesis of selected exemplar screening compounds is also described. In five cases, libraries were subsequently nominated for production on the basis of the scope and limitations of the validation work, as well as predicted molecular properties. In total, the research has led to the successful synthesis of >2500 novel alkaloid-like compounds for addition to the screening collection (the Joint European Compound Library, JECL) of the European Lead Factory. PMID:25600406

  13. Toxocara canis: Molecular basis of immune recognition and evasion

    PubMed Central

    Maizels, Rick M.

    2013-01-01

    Toxocara canis has extraordinary abilities to survive for many years in the tissues of diverse vertebrate species, as well as to develop to maturity in the intestinal tract of its definitive canid host. Human disease is caused by larval stages invading musculature, brain and the eye, and immune mechanisms appear to be ineffective at eliminating the infection. Survival of T. canis larvae can be attributed to two molecular strategies evolved by the parasite. Firstly, it releases quantities of ‘excretory–secretory’ products which include lectins, mucins and enzymes that interact with and modulate host immunity. For example, one lectin (CTL-1) is very similar to mammalian lectins, required for tissue inflammation, suggesting that T. canis may interfere with leucocyte extravasation into infected sites. The second strategy is the elaboration of a specialised mucin-rich surface coat; this is loosely attached to the parasite epicuticle in a fashion that permits rapid escape when host antibodies and cells adhere, resulting in an inflammatory reaction around a newly vacated focus. The mucins have been characterised as bearing multiple glycan side-chains, consisting of a blood-group-like trisaccharide with one or two O-methylation modifications. Both the lectins and these trisaccharides are targeted by host antibodies, with anti-lectin antibodies showing particular diagnostic promise. Antibodies to the mono-methylated trisaccharide appear to be T. canis-specific, as this epitope is not found in the closely related Toxocara cati, but all other antigenic determinants are very similar between the two species. This distinction may be important in designing new and more accurate diagnostic tests. Further tools to control toxocariasis could also arise from understanding the molecular cues and steps involved in larval development. In vitro-cultivated larvae express high levels of four mRNAs that are translationally silenced, as the proteins they encode are not detectable in

  14. Synthesis of molecularly imprinted polymer with 7-chloroethyl-theophylline-immobilized silica gel as template and its molecular recognition function

    NASA Astrophysics Data System (ADS)

    Zhang, Yuhui; Tong, Aijun; Li, Longdi

    2004-01-01

    By reaction of 7-chloroethyl-theophylline with aminopropylsilanized silica gel we synthesized a 7-chloroethyl-theophylline-immobilized silica gel as template molecule and prepared a molecularly imprinted polymer (MIP-Si), which had special recognition sites to 7-chloroethyl-theophylline. A conventional molecularly imprinted polymer (MIP) using 7-chloroethyl-theophylline as template was also prepared for comparison. Binding abilities to 7-chloroethyl-theophylline and its structural analogs revealed that the MIP-Si shows much higher binding speed and much more binding capacity than the MIP does.

  15. Brittle cornea syndrome: recognition, molecular diagnosis and management

    PubMed Central

    2013-01-01

    Brittle cornea syndrome (BCS) is an autosomal recessive disorder characterised by extreme corneal thinning and fragility. Corneal rupture can therefore occur either spontaneously or following minimal trauma in affected patients. Two genes, ZNF469 and PRDM5, have now been identified, in which causative pathogenic mutations collectively account for the condition in nearly all patients with BCS ascertained to date. Therefore, effective molecular diagnosis is now available for affected patients, and those at risk of being heterozygous carriers for BCS. We have previously identified mutations in ZNF469 in 14 families (in addition to 6 reported by others in the literature), and in PRDM5 in 8 families (with 1 further family now published by others). Clinical features include extreme corneal thinning with rupture, high myopia, blue sclerae, deafness of mixed aetiology with hypercompliant tympanic membranes, and variable skeletal manifestations. Corneal rupture may be the presenting feature of BCS, and it is possible that this may be incorrectly attributed to non-accidental injury. Mainstays of management include the prevention of ocular rupture by provision of protective polycarbonate spectacles, careful monitoring of visual and auditory function, and assessment for skeletal complications such as developmental dysplasia of the hip. Effective management depends upon appropriate identification of affected individuals, which may be challenging given the phenotypic overlap of BCS with other connective tissue disorders. PMID:23642083

  16. Molecular Recognition: Use of Metal-Containing Molecular Clefts for Supramolecular Self-Assembly and Host-Guest Formation

    SciTech Connect

    Crowley, James D.; Bosnich, Brice

    2008-10-03

    Molecular clefts consisting of a rigid spacer linked to two parallel cofacially disposed terpy-M-X (M = Pd{sup 2+}, Pt{sup 2+}) units, which can vary in separation from 6.6 to 7.2 {angstrom}, have been used as molecular receptors and for self-assembly with linear and triangular linkers to produce rectangles and trigonal prisms, respectively. Aromatic molecules form multiple host-guest adducts with the molecular cleft receptors and with the rectangles and trigonal prisms. Planar complexes of Pt{sup 2+} also form host-guest adducts. The forces that control this molecular recognition, namely, {pi}-{pi} interactions, charge-induced dipole interactions, charge-charge forces, weak metal-metal interactions and solvation effects, are discussed and assigned to the various adducts.

  17. Functional proteomic and structural insights into molecular recognition in the nitrilase family enzymes

    PubMed Central

    Barglow, Katherine T.; Saikatendu, Kumar S.; Bracey, Michael H.; Huey, Ruth; Morris, Garrett M.; Olson, Arthur J.; Stevens, Raymond C.; Cravatt, Benjamin F.

    2009-01-01

    Nitrilases are a large and diverse family of non-peptidic C-N hydrolases. The mammalian genome encodes eight nitrilase enzymes, several of which remain poorly characterized. Prominent among these are nitrilase-1 (Nit1) and nitrilase-2 (Nit2), which, despite having been shown to exert effects on cell growth and possibly serving as tumor suppressor genes, are without known substrates or selective inhibitors. In previous studies, we identified several nitrilases, including Nit1 and Nit2, as targets for dipeptide-chloroacetamide activity-based proteomics probes. Here, we have used these probes, in combination with high-resolution crystallography and molecular modeling, to systematically map the active site of Nit2 and identify residues involved in molecular recognition. We report the 1.4 Å crystal structure of mouse Nit2, and use this structure to identify residues that discriminate probe-labeling between the Nit1 and Nit2 enzymes. Interestingly, some of these residues are conserved across all vertebrate Nit2 enzymes and, conversely, not found in any vertebrate Nit1 enzymes, suggesting that they are key discriminators of molecular recognition between these otherwise highly homologous enzymes. Our findings thus point to a limited set of active site residues that establish distinct patterns of molecular recognition among nitrilases and provide chemical probes to selectively perturb the function of these enzymes in biological systems. PMID:19053248

  18. Molecular basis of sequence-specific recognition of pre-ribosomal RNA by nucleolin

    PubMed Central

    Allain, Frédéric H.-T.; Bouvet, Philippe; Dieckmann, Thorsten; Feigon, Juli

    2000-01-01

    The structure of the 28 kDa complex of the first two RNA binding domains (RBDs) of nucleolin (RBD12) with an RNA stem–loop that includes the nucleolin recognition element UCCCGA in the loop was determined by NMR spectroscopy. The structure of nucleolin RBD12 with the nucleolin recognition element (NRE) reveals that the two RBDs bind on opposite sides of the RNA loop, forming a molecular clamp that brings the 5′ and 3′ ends of the recognition sequence close together and stabilizing the stem–loop. The specific interactions observed in the structure explain the sequence specificity for the NRE sequence. Binding studies of mutant proteins and analysis of conserved residues support the proposed interactions. The mode of interaction of the protein with the RNA and the location of the putative NRE sites suggest that nucleolin may function as an RNA chaperone to prevent improper folding of the nascent pre-rRNA. PMID:11118222

  19. Molecular mechanisms of substrate recognition and specificity of botulinum neurotoxin serotype F.

    PubMed

    Chen, Sheng; Wan, Hoi Ying

    2011-01-15

    BoNTs (botulinum neurotoxins) are both deadly neurotoxins and natural toxins that are widely used in protein therapies to treat numerous neurological disorders of dystonia and spinal spasticity. Understanding the mechanism of action and substrate specificity of BoNTs is a prerequisite to develop antitoxin and novel BoNT-derived protein therapy. To date, there is a lack of detailed information with regard to how BoNTs recognize and hydrolyse the substrate VAMP-2 (vesicle-associated membrane protein 2), even though it is known to be cleaved by four of the seven BoNT serotypes, B, D, F, G and TeNT (tetanus neurotoxin). In the present study we dissected the molecular mechanisms of VAMP-2 recognition by BoNT serotype F for the first time. The initial substrate recognition was mediated through sequential binding of VAMP-2 to the B1, B2 and B3 pockets in LC/F (light chain of BoNT serotype F), which directed VAMP-2 to the active site of LC/F and stabilized the active site substrate recognition, where the P2, P1' and P2' sites of VAMP-2 were specifically recognized by the S2, S1' and S2' pockets of LC/F to promote substrate hydrolysis. The understanding of the molecular mechanisms of LC/F substrate recognition provides insights into the development of antitoxins and engineering novel BoNTs to optimize current therapy and extend therapeutic interventions. PMID:21029044

  20. Molecular recognition using corona phase complexes made of synthetic polymers adsorbed on carbon nanotubes

    NASA Astrophysics Data System (ADS)

    Zhang, Jingqing; Landry, Markita P.; Barone, Paul W.; Kim, Jong-Ho; Lin, Shangchao; Ulissi, Zachary W.; Lin, Dahua; Mu, Bin; Boghossian, Ardemis A.; Hilmer, Andrew J.; Rwei, Alina; Hinckley, Allison C.; Kruss, Sebastian; Shandell, Mia A.; Nair, Nitish; Blake, Steven; Şen, Fatih; Şen, Selda; Croy, Robert G.; Li, Deyu; Yum, Kyungsuk; Ahn, Jin-Ho; Jin, Hong; Heller, Daniel A.; Essigmann, John M.; Blankschtein, Daniel; Strano, Michael S.

    2013-12-01

    Understanding molecular recognition is of fundamental importance in applications such as therapeutics, chemical catalysis and sensor design. The most common recognition motifs involve biological macromolecules such as antibodies and aptamers. The key to biorecognition consists of a unique three-dimensional structure formed by a folded and constrained bioheteropolymer that creates a binding pocket, or an interface, able to recognize a specific molecule. Here, we show that synthetic heteropolymers, once constrained onto a single-walled carbon nanotube by chemical adsorption, also form a new corona phase that exhibits highly selective recognition for specific molecules. To prove the generality of this phenomenon, we report three examples of heteropolymer-nanotube recognition complexes for riboflavin, L-thyroxine and oestradiol. In each case, the recognition was predicted using a two-dimensional thermodynamic model of surface interactions in which the dissociation constants can be tuned by perturbing the chemical structure of the heteropolymer. Moreover, these complexes can be used as new types of spatiotemporal sensors based on modulation of the carbon nanotube photoemission in the near-infrared, as we show by tracking riboflavin diffusion in murine macrophages.

  1. Mode of molecular recognition of L-fucose by fucose-binding legume lectins.

    PubMed

    Thomas, C J; Surolia, A

    2000-02-16

    Recognition of cell surface carbohydrate moieties by lectins plays a vital role in many a biological process. Fucosyated residues are often implicated as key recognition markers in many cellular processes. In particular, the aspects of molecular recognition of fucose by fucose-bindinglectins UEA 1 and LTA pose a special case because no crystal structure of these lectins is available. The study was conducted to elucidate the process of recognition of l-fucose by UEA1 and LTA by correlating structure-based sequence alignment and other available biochemical/biophysical data. The study points out that the mode of recognition of l-fucose is coordinated by the invariant triad of residues the asparagine 137, glycine 105, and aspartate 87. The major hydrophobic stacking residue in this case is the tyrosine 220. The study also reiterates the key role of the conserved triad of residues in the combining site which is a common feature for all legume lectins whose crystal structures are known. PMID:10679191

  2. Aminoglycosides: Molecular Insights on the Recognition of RNA and Aminoglycoside Mimics

    PubMed Central

    Chittapragada, Maruthi; Roberts, Sarah; Ham, Young Wan

    2009-01-01

    RNA is increasingly recognized for its significant functions in biological systems and has recently become an important molecular target for therapeutics development. Aminoglycosides, a large class of clinically significant antibiotics, exert their biological functions by binding to prokaryotic ribosomal RNA (rRNA) and interfering with protein translation, resulting in bacterial cell death. They are also known to bind to viral mRNAs such as HIV-1 RRE and TAR. Consequently, aminoglycosides are accepted as the single most important model in understanding the principles that govern small molecule-RNA recognition, which is essential for the development of novel antibacterial, antiviral or even anti-oncogenic agents. This review outlines the chemical structures and mechanisms of molecular recognition and antibacterial activity of aminoglycosides and various aminoglycoside mimics that have recently been devised to improve biological efficacy, binding affinity and selectivity, or to circumvent bacterial resistance. PMID:19812740

  3. Magnetic deep eutectic solvents molecularly imprinted polymers for the selective recognition and separation of protein.

    PubMed

    Liu, Yanjin; Wang, Yuzhi; Dai, Qingzhou; Zhou, Yigang

    2016-09-14

    A novel and facile magnetic deep eutectic solvents (DES) molecularly imprinted polymers (MIPs) for the selective recognition and separation of Bovine hemoglobin (BHb) was prepared. The new-type DES was adopted as the functional monomer which would bring molecular imprinted technology to a new direction. The amounts of DES were optimized. The obtained magnetic DES-MIPs were characterized with fourier transform infrared spectrometry (FT-IR), thermogravimetric analysis (TGA), field emission scanning electron microscope (FESEM), dynamic light scattering (DLS), elemental analysis and vibrating sample magnetometer (VSM). The results suggested that the imprinted polymers were successfully formed and possessed a charming magnetism. The maximum adsorption capability (Qmax) and dissociation constant (KL) were analyzed by Langmuir isotherms (R(2) = 0.9983) and the value were estimated to be 175.44 mg/g and 0.035 mg/mL for the imprinted particles. And the imprinted particles showed a high imprinting factor of 4.77. In addition, the magnetic DES-MIPs presented outstanding recognition specificity and selectivity so that it can be utilized to separate template protein from the mixture of proteins and real samples. Last but not least, the combination of deep eutectic solvents and molecular imprinted technology in this paper provides a new perspective for the recognition and separation of proteins. PMID:27566352

  4. Organization of Inorganic Nanomaterials via Programmable DNA Self-Assembly and Peptide Molecular Recognition

    PubMed Central

    Carter, Joshua D.; LaBean, Thomas H.

    2011-01-01

    An interesting alternative to top-down nanofabrication is to imitate biology, where nanoscale materials frequently integrate organic molecules for self-assembly and molecular recognition with ordered, inorganic minerals to achieve mechanical, sensory, or other advantageous functions. Using biological systems as inspiration, researchers have sought to mimic the nanoscale composite materials produced in nature. Here, we describe a combination of self-assembly, molecular recognition, and templating, relying on an oligonucleotide covalently conjugated to a high-affinity gold-binding peptide. After integration of the peptide-coupled DNA into a self-assembling superstructure, the templated peptides recognize and bind gold nanoparticles. In addition to providing new ways of building functional multi-nanoparticle systems, this work provides experimental proof that a single peptide molecule is sufficient for immobilization of a nanoparticle. This molecular construction strategy, combining DNA assembly and peptide recognition, can be thought of as programmable, granular, artificial biomineralization. We also describe the important observation that the addition of 1–2% Tween 20 surfactant to the solution during gold particle binding allows the gold nanoparticles to remain soluble within the magnesium containing DNA assembly buffer under conditions that usually lead to the aggregation and precipitation of the nanoparticles. PMID:21314176

  5. Transfer of molecular recognition information from DNA nanostructures to gold nanoparticles.

    PubMed

    Edwardson, Thomas G W; Lau, Kai Lin; Bousmail, Danny; Serpell, Christopher J; Sleiman, Hanadi F

    2016-02-01

    DNA nanotechnology offers unparalleled precision and programmability for the bottom-up organization of materials. This approach relies on pre-assembling a DNA scaffold, typically containing hundreds of different strands, and using it to position functional components. A particularly attractive strategy is to employ DNA nanostructures not as permanent scaffolds, but as transient, reusable templates to transfer essential information to other materials. To our knowledge, this approach, akin to top-down lithography, has not been examined. Here we report a molecular printing strategy that chemically transfers a discrete pattern of DNA strands from a three-dimensional DNA structure to a gold nanoparticle. We show that the particles inherit the DNA sequence configuration encoded in the parent template with high fidelity. This provides control over the number of DNA strands and their relative placement, directionality and sequence asymmetry. Importantly, the nanoparticles produced exhibit the site-specific addressability of DNA nanostructures, and are promising components for energy, information and biomedical applications. PMID:26791900

  6. Combined Antiviral Therapy Using Designed Molecular Scaffolds Targeting Two Distinct Viral Functions, HIV-1 Genome Integration and Capsid Assembly

    PubMed Central

    Khamaikawin, Wannisa; Saoin, Somphot; Nangola, Sawitree; Chupradit, Koollawat; Sakkhachornphop, Supachai; Hadpech, Sudarat; Onlamoon, Nattawat; Ansari, Aftab A; Byrareddy, Siddappa N; Boulanger, Pierre; Hong, Saw-See; Torbett, Bruce E; Tayapiwatana, Chatchai

    2015-01-01

    Designed molecular scaffolds have been proposed as alternative therapeutic agents against HIV-1. The ankyrin repeat protein (AnkGAG1D4) and the zinc finger protein (2LTRZFP) have recently been characterized as intracellular antivirals, but these molecules, used individually, do not completely block HIV-1 replication and propagation. The capsid-binder AnkGAG1D4, which inhibits HIV-1 assembly, does not prevent the genome integration of newly incoming viruses. 2LTRZFP, designed to target the 2-LTR-circle junction of HIV-1 cDNA and block HIV-1 integration, would have no antiviral effect on HIV-1-infected cells. However, simultaneous expression of these two molecules should combine the advantage of preventive and curative treatments. To test this hypothesis, the genes encoding the N-myristoylated Myr(+)AnkGAG1D4 protein and the 2LTRZFP were introduced into human T-cells, using a third-generation lentiviral vector. SupT1 cells stably expressing 2LTRZFP alone or with Myr(+)AnkGAG1D4 showed a complete resistance to HIV-1 in viral challenge. Administration of the Myr(+)AnkGAG1D4 vector to HIV-1-preinfected SupT1 cells resulted in a significant antiviral effect. Resistance to viral infection was also observed in primary human CD4+ T-cells stably expressing Myr(+)AnkGAG1D4, and challenged with HIV-1, SIVmac, or SHIV. Our data suggest that our two anti-HIV-1 molecular scaffold prototypes are promising antiviral agents for anti-HIV-1 gene therapy. PMID:26305555

  7. Triplex molecular beacons for sensitive recognition of melamine based on abasic-site-containing DNA and fluorescent silver nanoclusters.

    PubMed

    Wang, Ya; Sun, Qianqian; Zhu, Linling; Zhang, Junying; Wang, Fengyang; Lu, Linlin; Yu, Haijun; Xu, Zhiai; Zhang, Wen

    2015-05-01

    A melamine aptamer derived from an abasic-site-containing triplex molecular beacon (tMB) was designed and developed for sensitive recognition of melamine by integrating tMBs and fluorescent silver nanoclusters (Ag NCs). PMID:25865656

  8. Hydrophilic Molecularly Imprinted Resorcinol-Formaldehyde-Melamine Resin Prepared in Water with Excellent Molecular Recognition in Aqueous Matrices.

    PubMed

    Lv, Tianwei; Yan, Hongyuan; Cao, Jiankun; Liang, Shiru

    2015-11-01

    Hydrophilic molecularly imprinted resorcinol-formaldehyde-melamine resin (MIRFM) is synthesized in water and shows excellent molecular recognition in aqueous matrices. The double functional monomers resorcinol and melamine, and the cross-linker formaldehyde, are all hydrophilic, and then the hydrophilic groups (such as hydroxyls, imino groups, and ether linkages) can be introduced into MIRFM, which make the material compatible with aqueous samples. The general principle is demonstrated by the synthesis of MIRFM using sulfanilamide as a dummy template for the selective recognition to sulfonamides (SAs) in milk samples. Resorcinol and melamine can interact with the template mainly by hydrogen bonding and π-π interaction, which makes MIRFM and the analytes have strong affinity. Besides, melamine can improve the rigidity of MIRFM and accelerate the polymerization process, so there is no need to add base or acid as a catalyst, which guarantees the success of molecular imprinting. MIRFM shows higher recovery and improved purification effect for SAs, in comparison to silica, HLB, C18, and SCX. Because of its excellent hydrophilicity and specificity, MIRFM is promising to be applied in biological, environmental, and clinical fields. PMID:26441379

  9. Molecular insights of protein contour recognition with ligand pharmacophoric sites through combinatorial library design and MD simulation in validating HTLV-1 PR inhibitors.

    PubMed

    Selvaraj, Chandrabose; Omer, Ankur; Singh, Poonam; Singh, Sanjeev Kumar

    2015-01-01

    Retroviruses HIV-1 and HTLV-1 are chiefly considered to be the most dangerous pathogens in Homo sapiens. These two viruses have structurally unique protease (PR) enzymes, which are having common function of its replication mechanism. Though HIV PR drugs failed to inhibit HTLV-1 infections, they emphatically emphasise the need for designing new lead compounds against HTLV-1 PR. Therefore, we tried to understand the binding level interactions through the charge environment present in both ligand and protein active sites. The domino effect illustrates that libraries of purvalanol-A are attuned to fill allosteric binding site of HTLV-1 PR through molecular recognition and shows proper binding of ligand pharmacophoric features in receptor contours. Our screening evaluates seven compounds from purvalanol-A libraries, and these compounds' pharmacophore searches for an appropriate place in the binding site and it places well according to respective receptor contour surfaces. Thus our result provides a platform for the progress of more effective compounds, which are better in free energy calculation, molecular docking, ADME and molecular dynamics studies. Finally, this research provided novel chemical scaffolds for HTLV-1 drug discovery. PMID:25335799

  10. Design Molecular Recognition Materials for Chiral Sensors, Separtations and Catalytic Materials

    SciTech Connect

    Jia, S.; Nenoff, T.M.; Provencio, P.; Qiu, Y.; Shelnutt, J.A.; Thoma, S.G.; Zhang, J.

    1998-11-01

    The goal is the development of materials that are highly sensitive and selective for chid chemicals and biochemical (such as insecticides, herbicides, proteins, and nerve agents) to be used as sensors, catalysts and separations membranes. Molecular modeling methods are being used to tailor chiral molecular recognition sites with high affinity and selectivity for specified agents. The work focuses on both silicate and non-silicate materials modified with chirally-pure fictional groups for the catalysis or separations of enantiomerically-pure molecules. Surfactant and quaternary amine templating is being used to synthesize porous frameworks, containing mesopores of 30 to 100 angstroms. Computer molecukw modeling methods are being used in the design of these materials, especially in the chid surface- modi~ing agents. Molecular modeling is also being used to predict the catalytic and separations selectivities of the modified mesoporous materials. The ability to design and synthesize tailored asymmetric molecular recognition sites for sensor coatings allows a broader range of chemicals to be sensed with the desired high sensitivity and selectivity. Initial experiments target the selective sensing of small molecule gases and non-toxic model neural compounds. Further efforts will address designing sensors that greatly extend the variety of resolvable chemical species and forming a predictive, model-based method for developing advanced sensors.

  11. Site-selective Characterization of Src Homology 3 Domain Molecular Recognition with Cyanophenylalanine Infrared Probes

    PubMed Central

    Horness, Rachel E.; Basom, Edward J.; Thielges, Megan C.

    2015-01-01

    Local heterogeneity of microenvironments in proteins is important in biological function, but difficult to characterize experimentally. One approach is the combination of infrared (IR) spectroscopy and site-selective incorporation of probe moieties with spectrally resolved IR absorptions that enable characterization within inherently congested protein IR spectra. We employed this method to study molecular recognition of a Src homology 3 (SH3) domain from the yeast protein Sho1 for a peptide containing the proline-rich recognition sequence of its physiological binding partner Pbs2. Nitrile IR probes were introduced at four distinct sites in the protein by selective incorporation of p-cyanophenylalanine via the amber codon suppressor method and characterized by IR spectroscopy. Variation among the IR absorption bands reports on heterogeneity in local residue environments dictated by the protein structure, as well as on residue-dependent changes upon peptide binding. The study informs on the molecular recognition of SH3Sho1 and illustrates the speed and simplicity of this approach for characterization of select microenvironments within proteins. PMID:26491469

  12. A molecular code dictates sequence-specific DNA recognition by homeodomains.

    PubMed Central

    Damante, G; Pellizzari, L; Esposito, G; Fogolari, F; Viglino, P; Fabbro, D; Tell, G; Formisano, S; Di Lauro, R

    1996-01-01

    Most homeodomains bind to DNA sequences containing the motif 5'-TAAT-3'. The homeodomain of thyroid transcription factor 1 (TTF-1HD) binds to sequences containing a 5'-CAAG-3' core motif, delineating a new mechanism for differential DNA recognition by homeodomains. We investigated the molecular basis of the DNA binding specificity of TTF-1HD by both structural and functional approaches. As already suggested by the three-dimensional structure of TTF-1HD, the DNA binding specificities of the TTF-1, Antennapedia and Engrailed homeodomains, either wild-type or mutants, indicated that the amino acid residue in position 54 is involved in the recognition of the nucleotide at the 3' end of the core motif 5'-NAAN-3'. The nucleotide at the 5' position of this core sequence is recognized by the amino acids located in position 6, 7 and 8 of the TTF-1 and Antennapedia homeodomains. These data, together with previous suggestions on the role of amino acids in position 50, indicate that the DNA binding specificity of homeodomains can be determined by a combinatorial molecular code. We also show that some specific combinations of the key amino acid residues involved in DNA recognition do not follow a simple, additive rule. Images PMID:8890172

  13. A norepinephrine coated magnetic molecularly imprinted polymer for simultaneous multiple chiral recognition.

    PubMed

    Chen, Juan; Liang, Ru-Ping; Wang, Xiao-Ni; Qiu, Jian-Ding

    2015-08-28

    A newly designed molecularly imprinted polymer (MIP) material was developed and successfully used as recognition element for enantioselective recognition by microchip electrophoresis. In this work, molecularly imprinted polymers were facilely prepared employing Fe3O4 nanoparticles (NPs) as the supporting substrate and norepinephrine as the functional monomer in the presence of template molecule in a weak alkaline solution. After extracting the embedded template molecules, the produced imprinted Fe3O4@polynorepinephrine (MIP-Fe3O4@PNE) NPs have cavities complementary to three dimensional shape of template molecules favoring high binding capacity and magnetism property for easy manipulation. The MIP-Fe3O4@PNE NPs prepared with l-tryptophan, l-valine, l-threonine, Gly-l-Phe, S-(-)-ofloxacin or S-(-)-binaphthol as template molecules were packed in the polydimethylsiloxane microchannel via magnetic field as novel stationary phase to successful enantioseparation of corresponding target analysts. The MIP-Fe3O4@PNE NPs-based microchip electrophoresis system exhibited strong recognition ability, excellent high-performance, admirable reproducibility and stability, which provided a powerful protocol for separation enantiomers within a short analytical time and opened up an avenue for multiplex chiral compound assay in various systems. PMID:26206627

  14. Conformational diversity of bacterial FabH: implications for molecular recognition specificity.

    PubMed

    Mittal, Anuradha; Johnson, Michael E

    2015-02-01

    The molecular basis of variable substrate and inhibitor specificity of the highly conserved bacterial fatty acid synthase enzyme, FabH, across different bacterial species remains poorly understood. In the current work, we explored the conformational diversity of FabH enzymes to understand the determinants of diverse interaction specificity across Gram-positive and Gram-negative bacteria. Atomistic molecular dynamics simulations reveal that FabH from E. coli and E. faecalis exhibit distinct native state conformational ensembles and dynamic behaviors. Despite strikingly similar substrate binding pockets, hot spot assessment using computational solvent mapping identified quite different favorable binding interactions between the two homologs. Our data suggest that FabH utilizes protein dynamics and seemingly minor sequence and structural differences to modulate its molecular recognition and substrate specificity across bacterial species. These insights will potentially facilitate the rational design and development of antibacterial inhibitors against FabH enzymes. PMID:25437098

  15. Molecular Recognition in Glycolaldehyde, the Simplest Sugar: Two Isolated Hydrogen Bonds Win Over One Cooperative Pair

    PubMed Central

    Altnöder, Jonas; Lee, Juhyon J; Otto, Katharina E; Suhm, Martin A

    2012-01-01

    Carbohydrates are used in nature as molecular recognition tools. Understanding their conformational behavior upon aggregation helps in rationalizing the way in which cells and bacteria use sugars to communicate. Here, the simplest α-hydroxy carbonyl compound, glycolaldehyde, was used as a model system. It was shown to form compact polar C2-symmetric dimers with intermolecular O–H⋅⋅⋅O=C bonds, while sacrificing the corresponding intramolecular hydrogen bonds. Supersonic jet infrared (IR) and Raman spectra combined with high-level quantum chemical calculations provide a consistent picture for the preference over more typical hydrogen bond insertion and addition patterns. Experimental evidence for at least one metastable dimer is presented. A rotational spectroscopy investigation of these dimers is encouraged, also in view of astrophysical searches. The binding motif competition of aldehydic sugars might play a role in chirality recognition phenomena of more complex derivatives in the gas phase. PMID:24551516

  16. Selective Nitrate Recognition by a Halogen-Bonding Four-Station [3]Rotaxane Molecular Shuttle.

    PubMed

    Barendt, Timothy A; Docker, Andrew; Marques, Igor; Félix, Vítor; Beer, Paul D

    2016-09-01

    The synthesis of the first halogen bonding [3]rotaxane host system containing a bis-iodo triazolium-bis-naphthalene diimide four station axle component is reported. Proton NMR anion binding titration experiments revealed the halogen bonding rotaxane is selective for nitrate over the more basic acetate, hydrogen carbonate and dihydrogen phosphate oxoanions and chloride, and exhibits enhanced recognition of anions relative to a hydrogen bonding analogue. This elaborate interlocked anion receptor functions via a novel dynamic pincer mechanism where upon nitrate anion binding, both macrocycles shuttle from the naphthalene diimide stations at the periphery of the axle to the central halogen bonding iodo-triazolium station anion recognition sites to form a unique 1:1 stoichiometric nitrate anion-rotaxane sandwich complex. Molecular dynamics simulations carried out on the nitrate and chloride halogen bonding [3]rotaxane complexes corroborate the (1) H NMR anion binding results. PMID:27436297

  17. Molecular recognition of parathyroid hormone by its G protein-coupled receptor

    SciTech Connect

    Pioszak, Augen A.; Xu, H. Eric

    2008-08-07

    Parathyroid hormone (PTH) is central to calcium homeostasis and bone maintenance in vertebrates, and as such it has been used for treating osteoporosis. It acts primarily by binding to its receptor, PTH1R, a member of the class B G protein-coupled receptor (GPCR) family that also includes receptors for glucagon, calcitonin, and other therapeutically important peptide hormones. Despite considerable interest and much research, determining the structure of the receptor-hormone complex has been hindered by difficulties in purifying the receptor and obtaining diffraction-quality crystals. Here, we present a method for expression and purification of the extracellular domain (ECD) of human PTH1R engineered as a maltose-binding protein (MBP) fusion that readily crystallizes. The 1.95-{angstrom} structure of PTH bound to the MBP-PTH1R-ECD fusion reveals that PTH docks as an amphipathic helix into a central hydrophobic groove formed by a three-layer {alpha}-{beta}-{beta}{alpha} fold of the PTH1R ECD, resembling a hot dog in a bun. Conservation in the ECD scaffold and the helical structure of peptide hormones emphasizes this hot dog model as a general mechanism of hormone recognition common to class B GPCRs. Our findings reveal critical insights into PTH actions and provide a rational template for drug design that targets this hormone signaling pathway.

  18. Molecular recognition with nanostructures fabricated by photopolymerization within metallic subwavelength apertures

    NASA Astrophysics Data System (ADS)

    Urraca, J. L.; Barrios, C. A.; Canalejas-Tejero, V.; Orellana, G.; Moreno-Bondi, M. C.

    2014-07-01

    The first demonstration of fabrication of submicron lateral resolution molecularly imprinted polymer (MIP) patterns by photoinduced local polymerization within metal subwavelength apertures is reported. The size of the photopolymerized MIP features is finely tuned by the dose of 532 nm radiation. Rhodamine 123 (R123) has been selected as a fluorescent model template to prove the recognition capability of the MIP nanostructures, which has been evaluated by fluorescence lifetime imaging microscopy (FLIM) with single photon timing measurements. The binding selectivity provided by the imprinting effect has been confirmed in the presence of compounds structurally related to R123. These results pave the way to the development of nanomaterial architectures with biomimetic artificial recognition properties for environmental, clinical and food testing.The first demonstration of fabrication of submicron lateral resolution molecularly imprinted polymer (MIP) patterns by photoinduced local polymerization within metal subwavelength apertures is reported. The size of the photopolymerized MIP features is finely tuned by the dose of 532 nm radiation. Rhodamine 123 (R123) has been selected as a fluorescent model template to prove the recognition capability of the MIP nanostructures, which has been evaluated by fluorescence lifetime imaging microscopy (FLIM) with single photon timing measurements. The binding selectivity provided by the imprinting effect has been confirmed in the presence of compounds structurally related to R123. These results pave the way to the development of nanomaterial architectures with biomimetic artificial recognition properties for environmental, clinical and food testing. Electronic supplementary information (ESI) available: Fig. SI.1: chemical structure and acronyms of the different fluorescent dyes; optimization of polymer composition; Table SI.1. Template recovery after polymerization; determination of the binding capacity by equilibrium rebinding

  19. Molecular recognition between glyconectins as an adhesion self-assembly pathway to multicellularity.

    PubMed

    Misevic, Gradimir N; Guerardel, Yann; Sumanovski, Lazar T; Slomianny, Marie-Christine; Demarty, Maurice; Ripoll, Camille; Karamanos, Yannis; Maes, Emmanuel; Popescu, Octavian; Strecker, Gerard

    2004-04-01

    The appearance of multicellular forms of life has been tightly coupled to the ability of an organism to retain its own anatomical integrity and to distinguish self from non-self. Large glycoconjugates, which make up the outermost cell surface layer of all Metazoans, are the primary candidates for the primordial adhesion and recognition functions in biological self-assembly systems. Atomic force microscopy experiments demonstrated that the binding strength between a single pair of Porifera cell surface glyconectin 1 glycoconjugates from Microciona prolifera can hold the weight of 1600 cells, proving their adhesion functions. Here, measurement of molecular self-recognition of glyconectins (GNs) purified from three Porifera species was used as an experimental model for primordial xenogeneic self/non-self discrimination. Physicochemical and biochemical characterization of the three glyconectins, their glycans, and peptides using gel electrophoresis, ultracentrifugation, NMR, mass spectrometry, glycosaminoglycan-degrading enzyme treatment, amino acid and carbohydrate analyses, and peptide mapping showed that GNs define a new family of proteoglycan-like molecules exhibiting species-specific structures with complex and repetitive acidic carbohydrate motives different from the classical proteoglycans and mucins. In functional self-assembly color-coded bead, cell, and blotting assays, glyconectins displayed species-specific recognition and adhesion. Affinity-purified monospecific polyclonal antibodies prepared against GN1, -2, and -3 glycans selectively inhibited cell adhesion of the respective sponge species. These results together with species-specific coaggregation of GN carbohydrate-coated beads with cells showed that GN glycans are functional in cell recognition and adhesion. The specificity of carbohydrate-mediated homophilic GN interactions in Porifera approaches the binding selectivity of the evolutionarily advanced immunoglobulin superfamily. Xenoselectivity of

  20. ADAPT, a Novel Scaffold Protein-Based Probe for Radionuclide Imaging of Molecular Targets That Are Expressed in Disseminated Cancers.

    PubMed

    Garousi, Javad; Lindbo, Sarah; Nilvebrant, Johan; Åstrand, Mikael; Buijs, Jos; Sandström, Mattias; Honarvar, Hadis; Orlova, Anna; Tolmachev, Vladimir; Hober, Sophia

    2015-10-15

    Small engineered scaffold proteins have attracted attention as probes for radionuclide-based molecular imaging. One class of these imaging probes, termed ABD-Derived Affinity Proteins (ADAPT), has been created using the albumin-binding domain (ABD) of streptococcal protein G as a stable protein scaffold. In this study, we report the development of a clinical lead probe termed ADAPT6 that binds HER2, an oncoprotein overexpressed in many breast cancers that serves as a theranostic biomarker for several approved targeting therapies. Surface-exposed amino acids of ABD were randomized to create a combinatorial library enabling selection of high-affinity binders to various proteins. Furthermore, ABD was engineered to enable rapid purification, to eradicate its binding to albumin, and to enable rapid blood clearance. Incorporation of a unique cysteine allowed site-specific conjugation to a maleimido derivative of a DOTA chelator, enabling radionuclide labeling, ¹¹¹In for SPECT imaging and ⁶⁸Ga for PET imaging. Pharmacologic studies in mice demonstrated that the fully engineered molecule (111)In/⁶⁸Ga-DOTA-(HE)3-ADAPT6 was specifically bound and taken up by HER2-expressing tumors, with a high tumor-to-normal tissue ratio in xenograft models of human cancer. Unbound tracer underwent rapid renal clearance followed by high renal reabsorption. HER2-expressing xenografts were visualized by gamma-camera or PET at 1 hour after infusion. PET experiments demonstrated feasibility for discrimination of xenografts with high or low HER2 expression. Our results offer a preclinical proof of concept for the use of ADAPT probes for noninvasive in vivo imaging. PMID:26297736

  1. Biophysical exploration of protein-flavonol recognition: effects of molecular properties and conformational flexibility.

    PubMed

    Ding, Fei; Peng, Wei; Peng, Yu-Kui

    2016-04-28

    The current work explores the biomolecular recognition of a series of flavonols by a protein and then uncovers the influences of the structural features of flavonols and the protein's own characteristics, e.g. the dynamics and flexibility, on the bioavailability of flavonols by using the pivotal biomacromolecule hemoglobin as a model. The experimental results revealed that flavonol may lead to a notable decrease in the steady-state fluorescence intensity of the β-37 Trp residue, and in the meantime the R-T transition of the protein transpired. Such noncovalent recognition forms the ground-state adduct, with an association intensity of 3.991 × 10(4) M(-1) in the reaction process, which has already been authenticated by the detailed analysis of time-resolved fluorescence and UV/vis absorption spectra. Furthermore, flavonol can form hydrogen bonds and π-conjugation effects with several amino acid residues on the polypeptide chain, for example, Trp-37, Arg-40, Asp-99 and Asn-102, and this event would induce self-regulation of the compact, regular conformation of the protein to a certain extent, which explicitly corroborates the results of circular dichroism. According to the study of molecular docking and structure-activity relationships, we could see that the recognition capacities of the protein-flavonols are inversely interrelated with the C log P values of the flavonol molecules. Moreover, the properties of the substituents in the structural B-ring unit of flavonols, i.e. polarity, position and number, will also prominently affect the degree of affinity and bioavailability of the protein-flavonol complexes. The analytical results of molecular dynamics (MD) simulation testified that the discussions of the structure-activity relationships are entirely logical, and the conformations of the amino acid residues forming noncovalent interactions tend to be stable in the MD simulation, as further elucidated from the dynamics data. Plainly, molecular recognition of

  2. Self-assembly of [3]catenanes and a [4]molecular necklace based on a cryptand/paraquat recognition motif.

    PubMed

    Ye, Yang; Wang, Shu-Ping; Zhu, Bin; Cook, Timothy R; Wu, Jing; Li, Shijun; Stang, Peter J

    2015-06-01

    Hierarchical self-assembly centered on metallacyclic scaffolds greatly facilitates the construction of mechanically interlocked structures. The formation of two [3]catenanes and one [4]molecular necklace is presented by utilizing the orthogonality of coordination-driven self-assembly and crown ether-based cryptand/paraquat derivative complexation. The threaded [3]catenanes and [4]molecular necklace were fabricated by using ten and nine total molecular components, respectively, from four and three unique species in solution, respectively. In all cases single supramolecular ensembles were obtained, attesting to the high degree of structural complexity made possible via self-assembly approaches. PMID:25996900

  3. Exploiting β-cyclodextrin in molecular imprinting for achieving recognition of benzylparaben in aqueous media.

    PubMed

    Asman, Saliza; Mohamad, Sharifah; Sarih, Norazilawati Muhamad

    2015-01-01

    The molecularly imprinted polymer (MIP) based on methacrylic acid functionalized β-cyclodextrin (MAA-β-CD) monomer was synthesized for the purpose of selective recognition of benzylparaben (BzP). The MAA-β-CD monomer was produced by bridging a methacrylic acid (MAA) and β-cyclodextrin (β-CD) using toluene-2,4-diisocyanate (TDI) by reacting the -OH group of MAA and one of the primary -OH groups of β-CD. This monomer comprised of triple interactions that included an inclusion complex, π-π interaction, and hydrogen bonding. To demonstrate β-CD performance in MIPs, two MIPs were prepared; molecularly imprinted polymer-methacrylic acid functionalized β-cyclodextrin, MIP(MAA-β-CD), and molecularly imprinted polymer-methacrylic acid, MIP(MAA); both prepared by a reversible addition fragmentation chain transfer polymerization (RAFT) in the bulk polymerization process. Both MIPs were characterized using the Fourier Transform Infrared Spectroscopy (FTIR), Field Emission Scanning Electron Microscopy (FESEM), and Brunauer-Emmett-Teller (BET). The presence of β-CD not only influenced the morphological structure, it also affected the specific surface area, average pore diameter, and total pore volume of the MIP. The rebinding of the imprinting effect was evaluated in binding experiments, which proved that the β-CD contributed significantly to the enhancement of the recognition affinity and selective adsorption of the MIP. PMID:25667978

  4. Probing Molecular Recognition at the Solid-Gas Interface by Sum-Frequency Vibrational Spectroscopy.

    PubMed

    Aprile, Arianna; Ciuchi, Federica; Pinalli, Roberta; Dalcanale, Enrico; Pagliusi, Pasquale

    2016-08-01

    Molecular recognition is among the most important chemical events in living systems and has been emulated in supramolecular chemistry, driven by chemical and biochemical sensing potential. Identifying host-guest association in situ at the interface, between the substrate-bound receptors and the analyte-containing media, is essential to predict complexation performances in term of the receptor conformation, orientation and organization. Herein, we report the first sum-frequency vibrational spectroscopy study of molecular recognition at the solid-gas interface. The binding capability of tetraquinoxaline cavitands toward volatile aromatic and aliphatic compounds, namely benzonitrile and acetonitrile, is investigated as test system. We prove the selective complexation of the receptors, organized in a solid-supported hybrid bilayer, toward aromatic compounds. Quantitative analysis allows to correlate the average orientations of the guest molecules and the host binding pockets, establishing "on-axis" complexation of benzonitrile within the cavitand cavity. The study is readily applicable to other receptors, molecular architectures, interfaces and analytes. PMID:27438350

  5. Improvement of DNA recognition through molecular imprinting: hybrid oligomer imprinted polymeric nanoparticles (oligoMIP NPs).

    PubMed

    Brahmbhatt, H; Poma, A; Pendergraff, H M; Watts, J K; Turner, N W

    2016-02-01

    High affinity and specific binding are cardinal properties of nucleic acids in relation to their biological function and their role in biotechnology. To this end, structural preorganization of oligonucleotides can significantly improve their binding performance, and numerous examples of this can be found in Nature as well as in artificial systems. Here we describe the production and characterization of hybrid DNA-polymer nanoparticles (oligoMIP NPs) as a system in which we have preorganized the oligonucleotide binding by molecular imprinting technology. Molecularly imprinted polymers (MIPs) are cost-effective "smart" polymeric materials capable of antibody-like detection, but characterized by superior robustness and the ability to work in extreme environmental conditions. Especially in the nanoparticle format, MIPs are dubbed as one of the most suitable alternatives to biological antibodies due to their selective molecular recognition properties, improved binding kinetics as well as size and dispersibility. Nonetheless, there have been very few attempts at DNA imprinting in the past due to structural complexity associated with these templates. By introducing modified thymine bases into the oligonucleotide sequences, which allow establishing covalent bonds between the DNA and the polymer, we demonstrate that such hybrid oligoMIP NPs specifically recognize their target DNA, and that the unique strategy of incorporating the complementary DNA strands as "preorganized selective monomers" improves the recognition properties without affecting the NPs physical properties such as size, shape or dispersibility. PMID:26509192

  6. Exploiting β-Cyclodextrin in Molecular Imprinting for Achieving Recognition of Benzylparaben in Aqueous Media

    PubMed Central

    Asman, Saliza; Mohamad, Sharifah; Muhamad Sarih, Norazilawati

    2015-01-01

    The molecularly imprinted polymer (MIP) based on methacrylic acid functionalized β-cyclodextrin (MAA-β-CD) monomer was synthesized for the purpose of selective recognition of benzylparaben (BzP). The MAA-β-CD monomer was produced by bridging a methacrylic acid (MAA) and β-cyclodextrin (β-CD) using toluene-2,4-diisocyanate (TDI) by reacting the –OH group of MAA and one of the primary –OH groups of β-CD. This monomer comprised of triple interactions that included an inclusion complex, π–π interaction, and hydrogen bonding. To demonstrate β-CD performance in MIPs, two MIPs were prepared; molecularly imprinted polymer-methacrylic acid functionalized β-cyclodextrin, MIP(MAA-β-CD), and molecularly imprinted polymer-methacrylic acid, MIP(MAA); both prepared by a reversible addition fragmentation chain transfer polymerization (RAFT) in the bulk polymerization process. Both MIPs were characterized using the Fourier Transform Infrared Spectroscopy (FTIR), Field Emission Scanning Electron Microscopy (FESEM), and Brunauer-Emmett-Teller (BET). The presence of β-CD not only influenced the morphological structure, it also affected the specific surface area, average pore diameter, and total pore volume of the MIP. The rebinding of the imprinting effect was evaluated in binding experiments, which proved that the β-CD contributed significantly to the enhancement of the recognition affinity and selective adsorption of the MIP. PMID:25667978

  7. Label-Free Sensing of Adenosine Based on Force Variations Induced by Molecular Recognition

    PubMed Central

    Li, Jingfeng; Li, Qing; Colombi Ciacchi, Lucio; Wei, Gang

    2015-01-01

    We demonstrate a simple force-based label-free strategy for the highly sensitive sensing of adenosine. An adenosine ssDNA aptamer was bound onto an atomic force microscopy (AFM) probe by covalent modification, and the molecular-interface adsorption force between the aptamer and a flat graphite surface was measured by single-molecule force spectroscopy (SMFS). In the presence of adenosine, the molecular recognition between adenosine and the aptamer resulted in the formation of a folded, hairpin-like DNA structure and hence caused a variation of the adsorption force at the graphite/water interface. The sensitive force response to molecular recognition provided an adenosine detection limit in the range of 0.1 to 1 nM. The addition of guanosine, cytidine, and uridine had no significant interference with the sensing of adenosine, indicating a strong selectivity of this sensor architecture. In addition, operational parameters that may affect the sensor, such as loading rate and solution ionic strength, were investigated. PMID:25808841

  8. Metal Oxide Nanosensors Using Polymeric Membranes, Enzymes and Antibody Receptors as Ion and Molecular Recognition Elements

    PubMed Central

    Willander, Magnus; Khun, Kimleang; Ibupoto, Zafar Hussain

    2014-01-01

    The concept of recognition and biofunctionality has attracted increasing interest in the fields of chemistry and material sciences. Advances in the field of nanotechnology for the synthesis of desired metal oxide nanostructures have provided a solid platform for the integration of nanoelectronic devices. These nanoelectronics-based devices have the ability to recognize molecular species of living organisms, and they have created the possibility for advanced chemical sensing functionalities with low limits of detection in the nanomolar range. In this review, various metal oxides, such as ZnO-, CuO-, and NiO-based nanosensors, are described using different methods (receptors) of functionalization for molecular and ion recognition. These functionalized metal oxide surfaces with a specific receptor involve either a complex formation between the receptor and the analyte or an electrostatic interaction during the chemical sensing of analytes. Metal oxide nanostructures are considered revolutionary nanomaterials that have a specific surface for the immobilization of biomolecules with much needed orientation, good conformation and enhanced biological activity which further improve the sensing properties of nanosensors. Metal oxide nanostructures are associated with certain unique optical, electrical and molecular characteristics in addition to unique functionalities and surface charge features which shows attractive platforms for interfacing biorecognition elements with effective transducing properties for signal amplification. There is a great opportunity in the near future for metal oxide nanostructure-based miniaturization and the development of engineering sensor devices. PMID:24841244

  9. Temperature-responsive molecular recognition chromatography using phenylalanine and tryptophan derived polymer modified silica beads.

    PubMed

    Hiruta, Yuki; Kanazashi, Ryosuke; Ayano, Eri; Okano, Teruo; Kanazawa, Hideko

    2016-02-01

    Temperature-responsive polymers incorporating molecular-recognition sites were developed as stationary phases for high-performance liquid chromatography (HPLC). The grafted stationary phases consisted of functional copolymers composed of N-isopropylacrylamide (NIPAAm) and N-acryloyl aromatic amino acid methyl esters, i.e., phenylalanine and tryptophan methyl esters (Phe-OMe and Trp-OMe). Three novel temperature-responsive polymers, P(NIPAAm-co-Phe-OMe5), P(NIPAAm-co-Phe-OMe10), and P(NIPAAm-co-Trp-OMe5), were synthesized. These copolymers exhibited a reversible hydrophilic/hydrophobic phase transition at their lower critical solution temperatures (LCSTs). The polymers were grafted onto aminopropyl silica using an activated ester-amine coupling method, and were packed into a stainless steel column, which was connected to an HPLC system. Temperature-responsive chromatography was conducted using water as the sole mobile phase. More hydrophobic analytes were retained longer, and the retention times of aromatic steroids and aromatic amino acids were dramatically increased. This indicated that π-π interactions occurred between the phenyl or indole moieties of phenylalanine or tryptophan, respectively, and the aromatic compounds. Furthermore, the retention times of compounds with hydrogen bond acceptors were higher with P(NIPAAm-co-Trp-OMe5), which contained indole as a hydrogen bond donor, than with P(NIPAAm-co-Phe-OMe5). This indicated that hydrogen bonding occurred between the stationary phase and the analytes. These results indicate that hydrophobic, π-π, and hydrogen bonding interactions all affected the separation mode of the temperature-responsive chromatography, and led to selective separation with molecular recognition. Both temperature-response and molecular recognition characteristics are present in the proposed separation system that utilizes a temperature-responsive polymer bearing aromatic amino acid derivatives. PMID:26646169

  10. Wiring of redox enzymes on three dimensional self-assembled molecular scaffold.

    PubMed

    Frasconi, Marco; Heyman, Arnon; Medalsy, Izhar; Porath, Danny; Mazzei, Franco; Shoseyov, Oded

    2011-10-18

    The integration of biological molecules and nanoscale components provides a fertile basis for the construction of hybrid materials of synergic properties and functions. Stable protein 1 (SP1), a highly stable ring shaped protein, was recently used to display different functional domains, to bind nanoparticles (NPs), and to spontaneously form two and three-dimensional structures. Here we show an approach to wire redox enzymes on this self-assembled protein-nanoparticle hybrid. Those hybrids are genetically engineered SP1s, displaying glucose oxidase (GOx) enzymes tethered to the protein inner pore. Moreover, the Au-NP-protein hybrids self-assembled to multiple enzymatic layers on the surface. By wiring the redox enzymes to the electrode, we present an active structure for the bioelectrocatalytic oxidation of glucose. This system demonstrates for the first time a three-dimensional assembly of multiple catalytic modules on a protein scaffold with an efficient electrical wiring of the enzyme units on an electrode surface, thus implementing a hybrid electrically active unit for nanobioelectronic applications. PMID:21895003

  11. In Vitro Selection of Cancer Cell-Specific Molecular Recognition Elements from Amino Acid Libraries

    PubMed Central

    Williams, Ryan M.; Sooter, Letha J.

    2015-01-01

    Differential cell systematic evolution of ligands by exponential enrichment (SELEX) is an in vitro selection method for obtaining molecular recognition elements (MREs) that specifically bind to individual cell types with high affinity. MREs are selected from initial large libraries of different nucleic or amino acids. This review outlines the construction of peptide and antibody fragment libraries as well as their different host types. Common methods of selection are also reviewed. Additionally, examples of cancer cell MREs are discussed, as well as their potential applications. PMID:26436100

  12. Raman spectroscopic protocol for the molecular recognition of key biomarkers in astrobiological exploration.

    PubMed

    Edwards, Howell G M

    2004-02-01

    Raman spectroscopy is proposed as novel instrumentation for the remote, robotic exploration of planetary surfaces, especially Mars. In recent years, information about the chemicals produced by organisms at the terrestrial limits of life, such as those surviving in Antarctic habitats, has facilitated the assembly of a spectral database of key biomarkers. In addition biogeological modifications which are essential for the survival strategies of environmentally stressed organisms have been identified. In this paper, the requirements for Raman spectroscopic instrumental detection of key bio--and bio-geological markers are outlined and a preliminary protocol established for the molecular spectral recognition of biological signatures in remote astrobiological exploration. PMID:14979640

  13. In Vitro Selection of a Single-Stranded DNA Molecular Recognition Element Specific for Bromacil

    PubMed Central

    Williams, Ryan M.; Kulick, Amanda R.; Yedlapalli, Srilakshmi; Battistella, Louisa; Hajiran, Cyrus J.; Sooter, Letha J.

    2014-01-01

    Bromacil is a widely used herbicide that is known to contaminate environmental systems. Due to the hazards it presents and inefficient detection methods, it is necessary to create a rapid and efficient sensing device. Towards this end, we have utilized a stringent in vitro selection method to identify single-stranded DNA molecular recognition elements (MRE) specific for bromacil. We have identified one MRE with high affinity (Kd = 9.6 nM) and specificity for bromacil compared to negative targets of selection and other pesticides. The selected ssDNA MRE will be useful as the sensing element in a field-deployable bromacil detection device. PMID:25400940

  14. An Elaborate Supramolecular Assembly for a Smart Nanodevice for Ratiometric Molecular Recognition and Logic Gates.

    PubMed

    Xie, Yu-Jie; Wu, Wen-Yu; Chen, Hao; Li, Xiang; Zhang, Hao-Li; Liu, Liang-Liang; Shao, Xing-Xin; Shan, Chang-Fu; Liu, Wei-Sheng; Tang, Yu

    2016-06-01

    Ingenious approaches to supramolecular assembly for fabricating smart nanodevices is one of the more significant topics in nanomaterials research. Herein, by using surface quaternized cationic carbon dots (CDots) as the assembly and fluorescence platform, anionic sulfonatocalix[4]arene with modifiable lower and upper rims as a connector, as well as in situ coordination of Tb(3+) ions, we propose an elaborate supramolecular assembly strategy for the facile fabrication of a multifunctional nanodevice. The dynamic equilibrium characteristics of the supramolecular interaction can eventually endow this nanodevice with functions of fluorescent ratiometric molecular recognition and as a nano-logic gate with two output channels. PMID:27106796

  15. Molecular Recognition Analyzed by Docking Simulations: The Aspartate Receptor and Isocitrate Dehydrogenase from Escherichia coli

    NASA Astrophysics Data System (ADS)

    Stoddard, Barry L.; Koshland, Daniel E., Jr.

    1993-02-01

    Protein docking protocols are used for the prediction of both small molecule binding to DNA and protein macromolecules and of complexes between macromolecules. These protocols are becoming increasingly automated and powerful tools for computer-aided drug design. We review the basic methodologies and strategies used for analyzing molecular recognition by computer docking algorithms and discuss recent experiments in which (i) substrate and substrate analogues are docked to the active site of isocitrate dehydrogenase and (ii) maltose binding protein is docked to the extracellular domain of the receptor, which signals maltose chemotaxis.

  16. Chiral recognition in adrenergic receptor binding mimics prepared by molecular imprinting.

    PubMed

    Ramström, O; Yu, C; Mosbach, K

    1996-01-01

    Molecularly imprinted polymers were prepared against the adrenomimetic agents ephedrine and pseudoephedrine. These compounds each incorporate two chiral centres. The polymers were evaluated with respect to enantiodiscrimination of various adrenergic ligands. The selectivity of the polymeric binding sites for the imprinted molecules was very high, and it was found that binding of both the enantiomeric and diastereomeric isomers of the imprint species were effectively obstructed, it was found that these polymers could selectively recognize the enantiomers of the endogenous adrenergic ligand epinephrine as well as several beta-adrenergic blockers. These observations suggest that these polymers effectively mimic the recognition patterns exhibited by natural adrenergic receptors. PMID:9174958

  17. Unprecedented selectivity in molecular recognition of carbohydrates by a metal-organic framework.

    PubMed

    Yabushita, Mizuho; Li, Peng; Bernales, Varinia; Kobayashi, Hirokazu; Fukuoka, Atsushi; Gagliardi, Laura; Farha, Omar K; Katz, Alexander

    2016-06-01

    Metal-organic framework (MOF) material NU-1000 adsorbs dimers cellobiose and lactose from aqueous solution, in amounts exceeding 1250 mg gNU-1000(-1) while completely excluding the adsorption of the monomer glucose, even in a competitive mode with cellobiose. The MOF also discriminates between dimers consisting of α and β linkages, showing no adsorption of maltose. Electronic structure calculations demonstrate that key to this selective molecular recognition is the number of favorable CH-π interactions made by the sugar with pyrene units of the MOF. PMID:27184781

  18. Molecular recognition of malachite green by hemoglobin and their specific interactions: insights from in silico docking and molecular spectroscopy.

    PubMed

    Peng, Wei; Ding, Fei; Peng, Yu-Kui; Sun, Ying

    2014-01-01

    Malachite green is an organic compound that can be widely used as a dyestuff for various materials; it has also emerged as a controversial agent in aquaculture. Since malachite green is proven to be carcinogenic and mutagenic, it may become a hazard to public health. For this reason, it is urgently required to analyze this controversial dye in more detail. In our current research, the interaction between malachite green and hemoglobin under physiological conditions was investigated by the methods of molecular modeling, fluorescence spectroscopy, circular dichroism (CD) as well as hydrophobic ANS displacement experiments. From the molecular docking, the central cavity of hemoglobin was assigned to possess high-affinity for malachite green, this result was corroborated by time-resolved fluorescence and hydrophobic ANS probe results. The recognition mechanism was found to be of static type, or rather the hemoglobin-malachite green complex formation occurred via noncovalent interactions such as π-π interactions, hydrogen bonds and hydrophobic interactions with an association constant of 10(4) M(-1). Moreover, the results also show that the spatial structure of the biopolymer was changed in the presence of malachite green with a decrease of the α-helix and increase of the β-sheet, turn and random coil suggesting protein damage, as derived from far-UV CD and three-dimensional fluorescence. Results of this work will help to further comprehend the molecular recognition of malachite green by the receptor protein and the possible toxicological profiles of other compounds, which are the metabolites and ramifications of malachite green. PMID:24226412

  19. Force-field development and molecular dynamics simulations of ferrocene-peptide conjugates as a scaffold for hydrogenase mimics

    SciTech Connect

    De Hatten, Xavier; Cournia, Zoe; Smith, Jeremy C; Huc, I; Metzler-Nolte, Nils

    2007-08-01

    The increasing importance of hydrogenase enzymes in the new energy research field has led us to examine the structure and dynamics of potential hydrogenase mimics, based on a ferrocene-peptide scaffold, using molecular dynamics (MD) simulations. To enable this MD study, a molecular mechanics force field for ferrocene-bearing peptides was developed and implemented in the CHARMM simulation package, thus extending the usefulness of the package into peptide-bioorganometallic chemistry. Using the automated frequency-matching method (AFMM), optimized intramolecular force-field parameters were generated through quantum chemical reference normal modes. The partial charges for ferrocene were derived by fitting point charges to quantum-chemically computed electrostatic potentials. The force field was tested against experimental X-ray crystal structures of dipeptide derivatives of ferrocene-1,1'-dicarboxylic acid. The calculations reproduce accurately the molecular geometries, including the characteristic C{sub 2}-symmetrical intramolecular hydrogen-bonding pattern, that were stable over 0.1 {micro}s MD simulations. The crystal packing properties of ferrocene-1-(D)alanine-(D)proline-1'-(D)alanine-(D)proline were also accurately reproduced. The lattice parameters of this crystal were conserved during a 0.1 {micro}s MD simulation and match the experimental values almost exactly. Simulations of the peptides in dichloromethane are also in good agreement with experimental NMR and circular dichroism (CD) data in solution. The developed force field was used to perform MD simulations on novel, as yet unsynthesized peptide fragments that surround the active site of [Ni-Fe] hydrogenase. The results of this simulation lead us to propose an improved design for synthetic peptide-based hydrogenase models. The presented MD simulation results of metallocenes thereby provide a convincing validation of our proposal to use ferrocene-peptides as minimal enzyme mimics.

  20. Force-field development and molecular dynamics simulations of ferrocene-peptide conjugates as a scaffold for hydrogenase mimics.

    SciTech Connect

    De Hatten, Xavier; Cournia, Zoe; Smith, Jeremy C; Metzler-Nolte, Nils

    2007-08-01

    The increasing importance of hydrogenase enzymes in the new energy research field has led us to examine the structure and dynamics of potential hydrogenase mimics, based on a ferrocene-peptide scaffold, using molecular dynamics (MD) simulations. To enable this MD study, a molecular mechanics force field for ferrocene-bearing peptides was developed and implemented in the CHARMM simulation package, thus extending the usefulness of the package into peptide-bioorganometallic chemistry. Using the automated frequency-matching method (AFMM), optimized intramolecular force-field parameters were generated through quantum chemical reference normal modes. The partial charges for ferrocene were derived by fitting point charges to quantum-chemically computed electrostatic potentials. The force field was tested against experimental X-ray crystal structures of dipeptide derivatives of ferrocene-1,1{prime}-dicarboxylic acid. The calculations reproduce accurately the molecular geometries, including the characteristic C2-symmetrical intramolecular hydrogen-bonding pattern, that were stable over 0.1{micro}s MD simulations. The crystal packing properties of ferrocene-1-(D)alanine-(D)proline{prime}-1-(D)alanine-(D)proline were also accurately reproduced. The lattice parameters of this crystal were conserved during a 0.1 s MD simulation and match the experimental values almost exactly. Simulations of the peptides in dichloromethane are also in good agreement with experimental NMR and circular dichroism (CD) data in solution. The developed force field was used to perform MD simulations on novel, as yet unsynthesized peptide fragments that surround the active site of [Ni-Fe] hydrogenase. The results of this simulation lead us to propose an improved design for synthetic peptide-based hydrogenase models. The presented MD simulation results of metallocenes thereby provide a convincing validation of our proposal to use ferrocene-peptides as minimal enzyme mimics.

  1. The ribosome as an optimal decoder: a lesson in molecular recognition

    NASA Astrophysics Data System (ADS)

    Tlusty, Tsvi; Savir, Yonatan

    2013-03-01

    The ribosome is a complex molecular machine that, in order to synthesize proteins, has to decode mRNAs by pairing their codons with matching tRNAs. Decoding is a major determinant of fitness and requires accurate and fast selection of correct tRNAs among many similar competitors. However, it is unclear whether the present ribosome, and in particular its large deformations during decoding, are the outcome of adaptation to its task as a decoder or the result of other constraints. Here, we derive the energy landscape that provides optimal discrimination between competing substrates, and thereby optimal tRNA decoding. We show that the measured landscape of the prokaryotic ribosome is indeed sculpted in this way. This suggests that conformational changes of the ribosome and tRNA during decoding are means to obtain an optimal decoder. Our analysis puts forward a generic mechanism that may be utilized by other ribosomes and other molecular recognition systems.

  2. Universal statistical fluctuations in thermodynamics and kinetics of single molecular recognition.

    PubMed

    Zheng, Xiliang; Wang, Jin

    2016-03-28

    We investigated the main universal statistical distributions of single molecular recognition. The distributions of the single molecule binding free energy spectrum or density of states were characterized in the ligand-receptor binding energy landscape. The analytical results are consistent with the microscopic molecular simulations. The free energy distribution of different binding modes or states for a single molecule ligand receptor pair is approximately Gaussian near the mean and exponential at the tail. The equilibrium constant of single molecule binding is log-normal distributed near the mean and power law distributed near the tail. Additionally, we found that the kinetics distribution of single molecule ligand binding can be characterized by log-normal around the mean and power law distribution near the tail. This distribution is caused by exploration of the underlying inhomogeneous free energy landscape. Different ligand-receptor binding complexes have the same universal form of distribution but differ in parameters. PMID:26947972

  3. A case for molecular recognition in nuclear separations: sulfate separation from nuclear wastes.

    PubMed

    Moyer, Bruce A; Custelcean, Radu; Hay, Benjamin P; Sessler, Jonathan L; Bowman-James, Kristin; Day, Victor W; Kang, Sung-Ok

    2013-04-01

    In this paper, we present the case for molecular-recognition approaches for sulfate removal from radioactive wastes via the use of anion-sequestering systems selective for sulfate, using either liquid-liquid extraction or crystallization. Potential benefits of removing sulfate from the waste include improved vitrification of the waste, reduced waste-form volume, and higher waste-form performance, all of which lead to potential cleanup schedule acceleration and cost savings. The need for sulfate removal from radioactive waste, especially legacy tank wastes stored at the Hanford site, is reviewed in detail and primarily relates to the low solubility of sulfate in borosilicate glass. Traditional methods applicable to the separation of sulfate from radioactive wastes are also reviewed, with the finding that currently no technology has been identified and successfully demonstrated to meet this need. Fundamental research in the authors' laboratories targeting sulfate as an important representative of the class of oxoanions is based on the hypothesis that designed receptors may provide the needed ability to recognize sulfate under highly competitive conditions, in particular where the nitrate anion concentration is high. Receptors that have been shown to have promising affinity for sulfate, either in extraction or in crystallization experiments, include hexaurea tripods, tetraamide macrocycles, cyclo[8]pyrroles, calixpyrroles, and self-assembled urea-lined cages. Good sulfate selectivity observed in the laboratory provides experimental support for the proposed molecular-recognition approach. PMID:23134587

  4. Molecular basis for bacterial peptidoglycan recognition by LysM domains

    PubMed Central

    Mesnage, Stéphane; Dellarole, Mariano; Baxter, Nicola J.; Rouget, Jean-Baptiste; Dimitrov, Jordan D.; Wang, Ning; Fujimoto, Yukari; Hounslow, Andrea M.; Lacroix-Desmazes, Sébastien; Fukase, Koichi; Foster, Simon J.; Williamson, Michael P.

    2014-01-01

    Carbohydrate recognition is essential for growth, cell adhesion and signalling in all living organisms. A highly conserved carbohydrate binding module, LysM, is found in proteins from viruses, bacteria, fungi, plants and mammals. LysM modules recognize polysaccharides containing N-acetylglucosamine (GlcNAc) residues including peptidoglycan, an essential component of the bacterial cell wall. However, the molecular mechanism underpinning LysM–peptidoglycan interactions remains unclear. Here we describe the molecular basis for peptidoglycan recognition by a multimodular LysM domain from AtlA, an autolysin involved in cell division in the opportunistic bacterial pathogen Enterococcus faecalis. We explore the contribution of individual modules to the binding, identify the peptidoglycan motif recognized, determine the structures of free and bound modules and reveal the residues involved in binding. Our results suggest that peptide stems modulate LysM binding to peptidoglycan. Using these results, we reveal how the LysM module recognizes the GlcNAc-X-GlcNAc motif present in polysaccharides across kingdoms. PMID:24978025

  5. Molecular Mechanism for Fungal Cell Wall Recognition by Rice Chitin Receptor OsCEBiP.

    PubMed

    Liu, Simiao; Wang, Jizong; Han, Zhifu; Gong, Xinqi; Zhang, Heqiao; Chai, Jijie

    2016-07-01

    Chitin is the major component of fungal cell wall and serves as a molecular pattern that can be recognized by the receptor OsCEBiP in rice, a lysine motif (LysM) receptor-like protein (RLP), to trigger immune responses. The molecular mechanisms underlying chitin recognition remain elusive. Here we report the crystal structures of the ectodomain of OsCEBiP (OsCEBiP-ECD) in free and chitin-bound forms. The structures reveal that OsCEBiP-ECD contains three tandem LysMs followed by a novel structure fold of cysteine-rich domain. The structures showed that chitin binding induces no striking conformational changes in OsCEBiP. Structural comparison among N-acetylglucosamine (NAG) oligomer-bound LysMs revealed a highly conserved recognition mechanism, which is expected to facilitate study of other LysM-containing proteins for their NAG binding. Modeling study showed that chitin induces OsCEBiP homodimerization in a "sliding mode". Our data provide insights into rice chitin receptor-mediated immunity triggered by fungal cell wall. PMID:27238968

  6. Molecular recognition of surface-immobilized carbohydrates by a synthetic lectin

    PubMed Central

    Rauschenberg, Melanie; Fritz, Eva-Corrina; Schulz, Christian; Kaufmann, Tobias

    2014-01-01

    Summary The molecular recognition of carbohydrates and proteins mediates a wide range of physiological processes and the development of synthetic carbohydrate receptors (“synthetic lectins”) constitutes a key advance in biomedical technology. In this article we report a synthetic lectin that selectively binds to carbohydrates immobilized in a molecular monolayer. Inspired by our previous work, we prepared a fluorescently labeled synthetic lectin consisting of a cyclic dimer of the tripeptide Cys-His-Cys, which forms spontaneously by air oxidation of the monomer. Amine-tethered derivatives of N-acetylneuraminic acid (NANA), β-D-galactose, β-D-glucose and α-D-mannose were microcontact printed on epoxide-terminated self-assembled monolayers. Successive prints resulted in simple microarrays of two carbohydrates. The selectivity of the synthetic lectin was investigated by incubation on the immobilized carbohydrates. Selective binding of the synthetic lectin to immobilized NANA and β-D-galactose was observed by fluorescence microscopy. The selectivity and affinity of the synthetic lectin was screened in competition experiments. In addition, the carbohydrate binding of the synthetic lectin was compared with the carbohydrate binding of the lectins concanavalin A and peanut agglutinin. It was found that the printed carbohydrates retain their characteristic selectivity towards the synthetic and natural lectins and that the recognition of synthetic and natural lectins is strictly orthogonal. PMID:24991289

  7. A Case for Molecular Recognition in Nuclear Separations: Sulfate Separation from Nuclear Wastes

    SciTech Connect

    Moyer, Bruce A; Custelcean, Radu; Hay, Benjamin; Sessler, Jonathan L.; Bowman-James, Kristin; Day, Victor W.; Kang, S.O.

    2013-01-01

    In this paper, we present the case for molecular-recognition approaches for sulfate removal from radioactive wastes via the use of anion-sequestering systems selective for sulfate, using either liquid liquid extraction or crystallization. Potential benefits of removing sulfate from the waste include improved vitrification of the waste, reduced waste-form volume, and higher waste-form performance, all of which lead to potential cleanup schedule acceleration and cost savings. The need for sulfate removal from radioactive waste, especially legacy tank wastes stored at the Hanford site, is reviewed in detail and primarily relates to the low solubility of sulfate in borosilicate glass. Traditional methods applicable to the separation of sulfate from radioactive wastes are also reviewed, with the finding that currently no technology has been identified and successfully demonstrated to meet this need. Fundamental research in the authors laboratories targeting sulfate as an important representative of the class of oxoanions is based on the hypothesis that designed receptors may provide the needed ability to recognize sulfate under highly competitive conditions, in particular where the nitrate anion concentration is high. Receptors that have been shown to have promising affinity for sulfate, either in extraction or in crystallization experiments, include hexaurea tripods, tetraamide macrocycles, cyclo[8]pyrroles, calixpyrroles, and self-assembled urea-lined cages. Good sulfate selectivity observed in the laboratory provides experimental support for the proposed molecular-recognition approach.

  8. Molecular recognition in the human immunodeficiency virus capsid and antiviral design.

    PubMed

    Bocanegra, Rebeca; Rodríguez-Huete, Alicia; Fuertes, Miguel Ángel; Del Álamo, Marta; Mateu, Mauricio G

    2012-11-01

    Many compounds able to interfere with HIV-1 infection have been identified; some 25 of them have been approved for clinical use. Current anti-HIV-1 therapy involves the use of drug cocktails, which reduces the probability of virus escape. However, many issues remain, including drug toxicity and the emergence of drug-resistant mutant viruses, even in treated patients. Therefore, there is a constant need for the development of new anti-HIV-1 agents targeting other molecules in the viral cycle. The capsid protein CA plays a key role in many molecular recognition events during HIV-1 morphogenesis and uncoating, and is eliciting increased interest as a promising target for antiviral intervention. This article provides a structure-based, integrated review on the CA-binding small molecules and peptides identified to date, and their effects on virus capsid assembly and stability, with emphasis on recent results not previously reviewed. As a complement, we present novel experimental results on the development and proof-of-concept application of a combinatorial approach to study molecular recognition in CA and its inhibition by peptide compounds. PMID:22728445

  9. Recognition of damage-associated molecular patterns related to nucleic acids during inflammation and vaccination

    PubMed Central

    Jounai, Nao; Kobiyama, Kouji; Takeshita, Fumihiko; Ishii, Ken J.

    2012-01-01

    All mammalian cells are equipped with large numbers of sensors for protection from various sorts of invaders, who, in turn, are equipped with molecules containing pathogen-associated molecular patterns (PAMPs). Once these sensors recognize non-self antigens containing PAMPs, various physiological responses including inflammation are induced to eliminate the pathogens. However, the host sometimes suffers from chronic infection or continuous injuries, resulting in production of self-molecules containing damage-associated molecular patterns (DAMPs). DAMPs are also responsible for the elimination of pathogens, but promiscuous recognition of DAMPs through sensors against PAMPs has been reported. Accumulation of DAMPs leads to massive inflammation and continuous production of DAMPs; that is, a vicious circle leading to the development of autoimmune disease. From a vaccinological point of view, the accurate recognition of both PAMPs and DAMPs is important for vaccine immunogenicity, because vaccine adjuvants are composed of several PAMPs and/or DAMPs, which are also associated with severe adverse events after vaccination. Here, we review as the roles of PAMPs and DAMPs upon infection with pathogens or inflammation, and the sensors responsible for recognizing them, as well as their relationship with the development of autoimmune disease or the immunogenicity of vaccines. PMID:23316484

  10. Enzymatic elucidation of haemocyanin from Kuruma shrimp Marsupenaeus japonicus and its molecular recognition mechanism towards pathogens.

    PubMed

    Sivakamavalli, Jeyachandran; Vaseeharan, Baskaralingam

    2015-01-01

    Haemocyanin is an important non-specific immune protein present in the hemolymph of invertebrates, which have the ability to recognize the microbial pathogens and trigger the innate immune system. In this study, we isolated and purified the haemocyanin using gel filtration chromatography and investigated its microbial recognition mechanism against the invading pathogens. Kuruma shrimp Marsupenaeus japonicus haemocyanin showed the single band with a molecular weight of 76 kDa on SDS-PAGE and its molecular mass was analysed through the MALDI. Pathogen recognition mechanism of M. japonicus haemocyanin was detected through bacterial agglutination, agglutination inhibition and prophenoloxidase activity. M. japonicus haemocyanin agglutinate all human blood RBC types and showed the bacterial agglutination against all tested Gram positive Staphylococcus aureus, Enterococcus faecalis and Bacillus subtilis and Gram negative Pseudomonas aeruginosa, Proteus vulgaris and Vibrio parahaemolyticus at the concentrations ranging from 30 to 50 μg/ml. Agglutination was inhibited by 50-200 mM of N-acetylneuraminic acid, a-D-glucose, D-galactose and D-xylose. Our results suggest that, 76 kDa subunit of M. japonicus haemocyanin recognize the pathogenic surface proteins which are present on the outer membrane of the bacteria and mediates the bacterial agglutination through haemocytes. This bacterial agglutination was visualized through Confocal Laser Scanning Microscopy (CLSM). This present study would be helpful to explore the importance of haemocyanin in innate immune response of M. japonicus and its eliciting pathogen recognition mechanism leads to the development of innate immunity in crustaceans. PMID:25204648

  11. Novel High-Viscosity Polyacrylamidated Chitosan for Neural Tissue Engineering: Fabrication of Anisotropic Neurodurable Scaffold via Molecular Disposition of Persulfate-Mediated Polymer Slicing and Complexation

    PubMed Central

    Kumar, Pradeep; Choonara, Yahya E.; du Toit, Lisa C.; Modi, Girish; Naidoo, Dinesh; Pillay, Viness

    2012-01-01

    Macroporous polyacrylamide-grafted-chitosan scaffolds for neural tissue engineering were fabricated with varied synthetic and viscosity profiles. A novel approach and mechanism was utilized for polyacrylamide grafting onto chitosan using potassium persulfate (KPS) mediated degradation of both polymers under a thermally controlled environment. Commercially available high molecular mass polyacrylamide was used instead of the acrylamide monomer for graft copolymerization. This grafting strategy yielded an enhanced grafting efficiency (GE = 92%), grafting ratio (GR = 263%), intrinsic viscosity (IV = 5.231 dL/g) and viscometric average molecular mass (MW = 1.63 × 106 Da) compared with known acrylamide that has a GE = 83%, GR = 178%, IV = 3.901 dL/g and MW = 1.22 × 106 Da. Image processing analysis of SEM images of the newly grafted neurodurable scaffold was undertaken based on the polymer-pore threshold. Attenuated Total Reflectance-FTIR spectral analyses in conjugation with DSC were used for the characterization and comparison of the newly grafted copolymers. Static Lattice Atomistic Simulations were employed to investigate and elucidate the copolymeric assembly and reaction mechanism by exploring the spatial disposition of chitosan and polyacrylamide with respect to the reactional profile of potassium persulfate. Interestingly, potassium persulfate, a peroxide, was found to play a dual role initially degrading the polymers—“polymer slicing”—thereby initiating the formation of free radicals and subsequently leading to synthesis of the high molecular mass polyacrylamide-grafted-chitosan (PAAm-g-CHT)—“polymer complexation”. Furthermore, the applicability of the uniquely grafted scaffold for neural tissue engineering was evaluated via PC12 neuronal cell seeding. The novel PAAm-g-CHT exhibited superior neurocompatibility in terms of cell infiltration owing to the anisotropic porous architecture, high molecular mass mediated robustness, superior

  12. Physical, Spatial, and Molecular Aspects of Extracellular Matrix of In Vivo Niches and Artificial Scaffolds Relevant to Stem Cells Research.

    PubMed

    Akhmanova, Maria; Osidak, Egor; Domogatsky, Sergey; Rodin, Sergey; Domogatskaya, Anna

    2015-01-01

    Extracellular matrix can influence stem cell choices, such as self-renewal, quiescence, migration, proliferation, phenotype maintenance, differentiation, or apoptosis. Three aspects of extracellular matrix were extensively studied during the last decade: physical properties, spatial presentation of adhesive epitopes, and molecular complexity. Over 15 different parameters have been shown to influence stem cell choices. Physical aspects include stiffness (or elasticity), viscoelasticity, pore size, porosity, amplitude and frequency of static and dynamic deformations applied to the matrix. Spatial aspects include scaffold dimensionality (2D or 3D) and thickness; cell polarity; area, shape, and microscale topography of cell adhesion surface; epitope concentration, epitope clustering characteristics (number of epitopes per cluster, spacing between epitopes within cluster, spacing between separate clusters, cluster patterns, and level of disorder in epitope arrangement), and nanotopography. Biochemical characteristics of natural extracellular matrix molecules regard diversity and structural complexity of matrix molecules, affinity and specificity of epitope interaction with cell receptors, role of non-affinity domains, complexity of supramolecular organization, and co-signaling by growth factors or matrix epitopes. Synergy between several matrix aspects enables stem cells to retain their function in vivo and may be a key to generation of long-term, robust, and effective in vitro stem cell culture systems. PMID:26351461

  13. Physical, Spatial, and Molecular Aspects of Extracellular Matrix of In Vivo Niches and Artificial Scaffolds Relevant to Stem Cells Research

    PubMed Central

    Akhmanova, Maria; Osidak, Egor; Domogatsky, Sergey; Rodin, Sergey; Domogatskaya, Anna

    2015-01-01

    Extracellular matrix can influence stem cell choices, such as self-renewal, quiescence, migration, proliferation, phenotype maintenance, differentiation, or apoptosis. Three aspects of extracellular matrix were extensively studied during the last decade: physical properties, spatial presentation of adhesive epitopes, and molecular complexity. Over 15 different parameters have been shown to influence stem cell choices. Physical aspects include stiffness (or elasticity), viscoelasticity, pore size, porosity, amplitude and frequency of static and dynamic deformations applied to the matrix. Spatial aspects include scaffold dimensionality (2D or 3D) and thickness; cell polarity; area, shape, and microscale topography of cell adhesion surface; epitope concentration, epitope clustering characteristics (number of epitopes per cluster, spacing between epitopes within cluster, spacing between separate clusters, cluster patterns, and level of disorder in epitope arrangement), and nanotopography. Biochemical characteristics of natural extracellular matrix molecules regard diversity and structural complexity of matrix molecules, affinity and specificity of epitope interaction with cell receptors, role of non-affinity domains, complexity of supramolecular organization, and co-signaling by growth factors or matrix epitopes. Synergy between several matrix aspects enables stem cells to retain their function in vivo and may be a key to generation of long-term, robust, and effective in vitro stem cell culture systems. PMID:26351461

  14. First example of a modular porphyrinoid assembly capable of stabilizing different metal ions in a single molecular scaffold.

    PubMed

    Murugavel, Muthuchamy; Reddy, R V Ramana; Dey, Dhananjay; Sankar, Jeyaraman

    2015-10-01

    We report the synthesis and characterization of porphyrin-corrole-porphyrin (Por-Cor-Por) hybrids directly linked at the meso-meso positions for the first time. The stability and solubility of the trimer are carefully balanced by adding electron-withdrawing substituents to the corrole ring and sterically bulky groups on the porphyrins. The new hybrids are capable of stabilizing more than one metal ion in a single molecular scaffold. The versatility of the triad has been demonstrated by successfully stabilizing homo- (Ni) and heterotrinuclear (Ni-Cu-Ni) coordination motifs. The solid-state structure of the NiPor-CuCor-PorNi hybrid was revealed by single-crystal X-ray diffraction studies. The Ni(II) porphyrins are significantly ruffled and tilted by 83° from the plane of corrole. The robustness of the synthesized hybrids was reflected in the electrochemical investigations and the redox behaviour of the hybrids show that the oxidation processes are mostly corrole-centred. In particular it is worth noting that the Por-Cor-Por hybrid can further be manipulated due to the presence of substituent-free meso-positions on both the terminals. PMID:26242294

  15. Molecular recognition and colorimetric detection of cholera toxin by poly(diacetylene) liposomes incorporating G{sub m1} ganglioside

    SciTech Connect

    Pan, J.J.; Charych, D.

    1997-03-19

    Molecular recognition sites on cell membranes serve as the main communication channels between the inside of a cell and its surroundings. Upon receptor binding, cellular messages such as ion channel opening or activation of enzymes are triggered. In this report, we demonstrate that artificial cell membranes made from conjugated lipid polymers (poly(diacetylene)) can, on a simple level, mimic membrane processes of molecular recognition and signal transduction. The ganglioside GM1 was incorporated into poly(diacetylene) liposomes. Molecular recognition of cholera toxin at the interface of the liposome resulted in a change of the membrane color due to conformational charges in the conjugated (ene-yne) polymer backbone. The `colored liposomes` might be used as simple colorimetric sensors for drug screening or as new tools to study membrane-membrane or membrane-receptor interactions. 21 refs., 3 figs.

  16. Using molecular recognition of beta-cyclodextrin to determine molecular weights of low-molecular-weight explosives by MALDI-TOF mass spectrometry.

    PubMed

    Zhang, Min; Shi, Zhen; Bai, Yinjuan; Gao, Yong; Hu, Rongzu; Zhao, Fenqi

    2006-02-01

    This study presents a novel method for determining the molecular weights of low molecular weight (MW) energetic compounds through their complexes of beta-cyclodextrin (beta-CD) and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) in a mass range of 500 to 1700 Da, avoiding matrix interference. The MWs of one composite explosive composed of 2,6-DNT, TNT, and RDX, one propellant with unknown components, and 14 single-compound explosives (RDX, HMX, 3,4-DNT, 2,6-DNT, 2,5-DNT, 2,4,6-TNT, TNAZ, DNI, BTTN, NG, TO, NTO, NP, and 662) were measured. The molecular recognition and inclusion behavior of beta-CD to energetic materials (EMs) were investigated. The results show that (1) the established method is sensitive, simple, accurate, and suitable for determining the MWs of low-MW single-compound explosives and energetic components in composite explosives and propellants; and (2) beta-CD has good inclusion and modular recognition abilities to the above EMs. PMID:16406809

  17. Design, Synthesis, and Molecular Docking Studies of a Conjugated Thiadiazole-Thiourea Scaffold as Antituberculosis Agents.

    PubMed

    Tatar, Esra; Karakuş, Sevgi; Küçükgüzel, Şükriye Güniz; Öktem Okullu, Sinem; Ünübol, Nihan; Kocagöz, Tanıl; De Clercq, Erik; Andrei, Graciela; Snoeck, Robert; Pannecouque, Christophe; Kalaycı, Sadık; Şahin, Fikrettin; Sriram, Dharmarajan; Yogeeswari, Perumal; Küçükgüzel, İlkay

    2016-01-01

    In view of the emergence and frequency of multidrug-resistant and extensively drug-resistant tuberculosis and consequences of acquired resistance to clinically used drugs, we undertook the design and synthesis of novel prototypes that possess the advantage of the two pharmacophores of thiourea and 1,3,4-thiadiazole in a single molecular backbone. Three compounds from our series were distinguished from the others by their promising activity profiles against Mycobacterium tuberculosis strain H37Rv. Compounds 11 and 19 were the most active representatives with minimum inhibitory concentration (MIC) values of 10.96 and 11.48 µM, respectively. Compound 15 was shown to inhibit M. tuberculosis strain H37Rv with an MIC value of 17.81 µM. Cytotoxicity results in the Vero cell line showed that these three derivatives had selectivity indices between 1.8 and 8.7. In order to rationalize the biological results of our compounds, molecular docking studies with the enoyl acyl carrier protein reductase (InhA) of M. tuberculosis were performed and compounds 11, 15, and 19 were found to have good docking scores in the range of -7.12 to -7.83 kcal/mol. PMID:27040623

  18. γ-Cyclodextrin capped silver nanoparticles for molecular recognition and enhancement of antibacterial activity of chloramphenicol.

    PubMed

    Gannimani, Ramesh; Ramesh, Muthusamy; Mtambo, Sphamandla; Pillay, Karen; Soliman, Mahmoud E; Govender, Patrick

    2016-04-01

    Computational studies were conducted to identify the favourable formation of the inclusion complex of chloramphenicol with cyclodextrins. The results of molecular docking and molecular dynamics predicted the strongest interaction of chloramphenicol with γ-cyclodextrin. Further, the inclusion complex of chloramphenicol with γ-cyclodextrin was experimentally prepared and a phenomenon of inclusion was verified by using different characterization techniques such as thermogravimetric analysis, differential scanning calorimetry, (1)H nuclear magnetic resonance (NMR) and two dimensional nuclear overhauser effect spectroscopy (NOESY) experiments. From these results it was concluded that γ-cyclodextrins could be an appropriate cyclodextrin polymer which can be used to functionalize chloramphenicol on the surface of silver nanoparticles. In addition, γ-cyclodextrin capped silver nanoparticles were synthesized and characterized using UV-visible spectroscopy, scanning electron microscopy (SEM), transmission electron microscopy (TEM), energy dispersive X-ray analysis (EDX), Fourier transform infrared spectroscopy (FTIR) and zeta potential analysis. Molecular recognition of chloramphenicol by these cyclodextrin capped silver nanoparticles was confirmed by surface enhanced raman spectroscopy (SERS) experiments. Synergistic antibacterial effect of chloramphenicol with γ-cyclodextrin capped silver nanoparticles was evaluated against Pseudomonas aeruginosa (ATCC 27853), Enterococcus faecalis (ATCC 5129), Klebsiella pneumoniae (ATCC 700603) and Staphylococcus aureus (ATCC 43300). The results from the antibacterial experiment were favourable thus allowing us to conclude that the approach of modifying organic drug molecules with cyclodextrin capped inorganic silver nanoparticles could help to enhance the antibacterial activity of them. PMID:26824520

  19. Preparation and characterization of monodisperse molecularly imprinted polymers for the recognition and enrichment of oleanolic acid.

    PubMed

    Tang, Zonggui; Liu, Changbin; Wang, Jing; Li, Hongmin; Ji, Yong; Wang, Guohong; Lu, Chunxia

    2016-04-01

    Monodisperse molecularly imprinted polymers for oleanolic acid were successfully prepared by a precipitation polymerization method using oleanolic acid as a template, methacrylic acid as a functional monomer, and divinylbenzene/ethylene glycol dimethacrylate as a crosslinker in a mixture of acetonitrile and ethanol (3:1, v/v). The imprinted polymers and nonimprinted polymers were characterized by using scanning electron microscopy, Fourier transform infrared spectroscopy, and thermogravimetric analysis. The resulting imprinted polymers had average diameters of 3.15 μm and monodispersity values of 1.024. The results clearly demonstrate that use of ethanol as a cosolvent is indeed exceedingly effective in promoting the dissolution of oleanolic acid and in obtaining uniform microspheres. Molecular recognition properties and binding capability to oleanolic acid were evaluated by adsorption testing, which indicated that the imprinted polymers displayed optimal binding performance with a maximum adsorption capacity of 17.3 mg/g and a binding saturation time of 80 min. Meanwhile, the produced imprinted polymers exhibited higher selectivity to oleanolic acid than that for ursolic acid and rhein. Herein, the studies can provide theoretical and experimental references for the oleanolic acid molecular imprinted system. PMID:27106769

  20. DNA aptamers are functional molecular recognition sensors in protic ionic liquids.

    PubMed

    Machado, Isabel; Özalp, Veli Cengiz; Rezabal, Elixabete; Schäfer, Thomas

    2014-09-01

    The function and structural changes of an AMP molecular aptamer beacon and its molecular recognition capacity for its target, adenosine monophosphate (AMP), was systematically explored in solution with a protic ionic liquid, ethylammonium nitrate (EAN). It could be proven that up to 2 M of EAN in TBS buffer, the AMP molecular aptamer beacon was still capable of recognizing AMP while also maintaining its specificity. The specificity was proven by using the guanosine monophosphate (GMP) as target; GMP is structurally similar to AMP but was not recognized by the aptamer. We also found that in highly concentrated EAN solutions the overall amount of double stranded DNA formed, as well as its respective thermal stability, diminished gradually, but surprisingly the hybridization rate (kh ) of single stranded DNA was significantly accelerated in the presence of EAN. The latter may have important implications in DNA technology for the design of biosensing and DNA-based nanodevices in nonconventional solvents, such as ionic liquids. PMID:25065686

  1. Preparation of a magnetic molecularly imprinted polymer for selective recognition of rhodamine B

    NASA Astrophysics Data System (ADS)

    Liu, Xiuying; Yu, Dan; Yu, Yingchao; Ji, Shujuan

    2014-11-01

    A novel magnetic molecularly imprinted polymer (MMIP) was developed as an adsorbent to selectively remove rhodamine B from real samples. The polymer was characterized by scanning electron microscopy, Fourier-transform infrared spectroscopy, and thermo-gravimetric analysis. Static adsorption, kinetic adsorption, and selective recognition experiments were also performed to investigate the specific adsorption equilibrium, kinetics, and selective recognition ability of the MMIP. The MMIPs had outstanding thermal stability, large adsorption capacity, and high competitive selectivity. When they were used as dispersed solid-phase extraction adsorbents in real samples, rhodamine B recovery was 79.97-81.88% and 75.56-79.74% in intra-day and inter-day reproducibility experiments with relative standard deviations lower than 2.62% and 4.28%, respectively. Extraction was optimized for yield and efficiency. Precision, accuracy, and linear working range were determined under optimal experimental conditions. The limits of detection and quantification were 1.05 and 3.49 μg L-1, respectively. These results suggest MMIPs may be used for determination of rhodamine B in real samples.

  2. Molecular Understanding of USP7 Substrate Recognition and C-Terminal Activation.

    PubMed

    Rougé, Lionel; Bainbridge, Travis W; Kwok, Michael; Tong, Raymond; Di Lello, Paola; Wertz, Ingrid E; Maurer, Till; Ernst, James A; Murray, Jeremy

    2016-08-01

    The deubiquitinating enzyme USP7 has a pivotal role in regulating the stability of proteins involved in fundamental cellular processes of normal biology and disease. Despite the importance of USP7, the mechanisms underlying substrate recognition and catalytic activation are poorly understood. Here we present structural, biochemical, and biophysical analyses elucidating the molecular mechanism by which the C-terminal 19 amino acids of USP7 (residues 1084-1102) enhance the ubiquitin cleavage activity of the deubiquitinase (DUB) domain. Our data demonstrate that the C-terminal peptide binds the activation cleft in the catalytic domain and stabilizes the catalytically competent conformation of USP7. Additional structures of longer fragments of USP7, as well as solution studies, provide insight into full-length USP7, the role of the UBL domains, and demonstrate that both substrate recognition and deubiquitinase activity are highly regulated by the catalytic and noncatalytic domains of USP7, a feature that could be essential for the proper function of multi-domain DUBs. PMID:27452404

  3. Electrospun Nanofibers from a Tricyanofuran-Based Molecular Switch for Colorimetric Recognition of Ammonia Gas.

    PubMed

    Khattab, Tawfik A; Abdelmoez, Sherif; Klapötke, Thomas M

    2016-03-14

    A chromophore based on tricyanofuran (TCF) with a hydrazone (H) recognition moiety was developed. Its molecular-switching performance is reversible and has differential sensitivity towards aqueous ammonia at comparable concentrations. Nanofibers were fabricated from the TCF-H chromophore by electrospinning. The film fabricated from these nanofibers functions as a solid-state optical chemosensor for probing ammonia vapor. Recognition of ammonia vapor occurs by proton transfer from the hydrazone fragment of the chromophore to the ammonia nitrogen atom and is facilitated by the strongly electron withdrawing TCF fragment. The TCF-H chromophore was added to a solution of poly(acrylic acid), which was electrospun to obtain a nanofibrous sensor device. The morphology of the nanofibrous sensor was determined by SEM, which showed that nanofibers with a diameter range of 200-450 nm formed a nonwoven mat. The resultant nanofibrous sensor showed very good sensitivity in ammonia-vapor detection. Furthermore, very good reversibility and short response time were also observed. PMID:26864701

  4. Molecular basis of synaptic vesicle cargo recognition by the endocytic sorting adaptor stonin 2.

    PubMed

    Jung, Nadja; Wienisch, Martin; Gu, Mingyu; Rand, James B; Müller, Sebastian L; Krause, Gerd; Jorgensen, Erik M; Klingauf, Jürgen; Haucke, Volker

    2007-12-31

    Synaptic transmission depends on clathrin-mediated recycling of synaptic vesicles (SVs). How select SV proteins are targeted for internalization has remained elusive. Stonins are evolutionarily conserved adaptors dedicated to endocytic sorting of the SV protein synaptotagmin. Our data identify the molecular determinants for recognition of synaptotagmin by stonin 2 or its Caenorhabditis elegans orthologue UNC-41B. The interaction involves the direct association of clusters of basic residues on the surface of the cytoplasmic domain of synaptotagmin 1 and a beta strand within the mu-homology domain of stonin 2. Mutation of K783, Y784, and E785 to alanine within this stonin 2 beta strand results in failure of the mutant stonin protein to associate with synaptotagmin, to accumulate at synapses, and to facilitate synaptotagmin internalization. Synaptotagmin-binding-defective UNC-41B is unable to rescue paralysis in C. elegans stonin mutant animals, suggesting that the mechanism of stonin-mediated SV cargo recognition is conserved from worms to mammals. PMID:18166656

  5. Ultrasound Molecular Imaging of the Breast Cancer Neovasculature using Engineered Fibronectin Scaffold Ligands: A Novel Class of Targeted Contrast Ultrasound Agent

    PubMed Central

    Abou-Elkacem, Lotfi; Wilson, Katheryne E.; Johnson, Sadie M.; Chowdhury, Sayan M.; Bachawal, Sunitha; Hackel, Benjamin J.; Tian, Lu; Willmann, Jürgen K.

    2016-01-01

    Molecularly-targeted microbubbles (MBs) are increasingly being recognized as promising contrast agents for oncological molecular imaging with ultrasound. With the detection and validation of new molecular imaging targets, novel binding ligands are needed that bind to molecular imaging targets with high affinity and specificity. In this study we assessed a novel class of potentially clinically translatable MBs using an engineered 10th type III domain of human-fibronectin (MB-FN3VEGFR2) scaffold-ligand to image VEGFR2 on the neovasculature of cancer. The in vitro binding of MB-FN3VEGFR2 to a soluble VEGFR2 was assessed by flow-cytometry (FACS) and binding to VEGFR2-expressing cells was assessed by flow-chamber cell attachment studies under flow shear stress conditions. In vivo binding of MB-FN3VEGFR2 was tested in a transgenic mouse model (FVB/N Tg(MMTV/PyMT634Mul) of breast cancer and control litter mates with normal mammary glands. In vitro FACS and flow-chamber cell attachment studies showed significantly (P<0.01) higher binding to VEGFR2 using MB-FN3VEGFR2 than control agents. In vivo ultrasound molecular imaging (USMI) studies using MB-FN3VEGFR2 demonstrated specific binding to VEGFR2 and was significantly higher (P<0.01) in breast cancer compared to normal breast tissue. Ex vivo immunofluorescence-analysis showed significantly (P<0.01) increased VEGFR2-expression in breast cancer compared to normal mammary tissue. Our results suggest that MBs coupled to FN3-scaffolds can be designed and used for USMI of breast cancer neoangiogenesis. Due to their small size, stability, solubility, the lack of glycosylation and disulfide bonds, FN3-scaffolds can be recombinantly produced with the advantage of generating small, high affinity ligands in a cost efficient way for USMI. PMID:27570547

  6. Ultrasound Molecular Imaging of the Breast Cancer Neovasculature using Engineered Fibronectin Scaffold Ligands: A Novel Class of Targeted Contrast Ultrasound Agent.

    PubMed

    Abou-Elkacem, Lotfi; Wilson, Katheryne E; Johnson, Sadie M; Chowdhury, Sayan M; Bachawal, Sunitha; Hackel, Benjamin J; Tian, Lu; Willmann, Jürgen K

    2016-01-01

    Molecularly-targeted microbubbles (MBs) are increasingly being recognized as promising contrast agents for oncological molecular imaging with ultrasound. With the detection and validation of new molecular imaging targets, novel binding ligands are needed that bind to molecular imaging targets with high affinity and specificity. In this study we assessed a novel class of potentially clinically translatable MBs using an engineered 10(th) type III domain of human-fibronectin (MB-FN3VEGFR2) scaffold-ligand to image VEGFR2 on the neovasculature of cancer. The in vitro binding of MB-FN3VEGFR2 to a soluble VEGFR2 was assessed by flow-cytometry (FACS) and binding to VEGFR2-expressing cells was assessed by flow-chamber cell attachment studies under flow shear stress conditions. In vivo binding of MB-FN3VEGFR2 was tested in a transgenic mouse model (FVB/N Tg(MMTV/PyMT634Mul) of breast cancer and control litter mates with normal mammary glands. In vitro FACS and flow-chamber cell attachment studies showed significantly (P<0.01) higher binding to VEGFR2 using MB-FN3VEGFR2 than control agents. In vivo ultrasound molecular imaging (USMI) studies using MB-FN3VEGFR2 demonstrated specific binding to VEGFR2 and was significantly higher (P<0.01) in breast cancer compared to normal breast tissue. Ex vivo immunofluorescence-analysis showed significantly (P<0.01) increased VEGFR2-expression in breast cancer compared to normal mammary tissue. Our results suggest that MBs coupled to FN3-scaffolds can be designed and used for USMI of breast cancer neoangiogenesis. Due to their small size, stability, solubility, the lack of glycosylation and disulfide bonds, FN3-scaffolds can be recombinantly produced with the advantage of generating small, high affinity ligands in a cost efficient way for USMI. PMID:27570547

  7. Emergent Molecular Recognition through Self-Assembly: Unexpected Selectivity for Hyaluronic Acid among Glycosaminoglycans.

    PubMed

    Noguchi, Takao; Roy, Bappaditya; Yoshihara, Daisuke; Sakamoto, Junji; Yamamoto, Tatsuhiro; Shinkai, Seiji

    2016-05-01

    Oligophenylenevinylene (OPV)-based fluorescent (FL) chemosensors exhibiting linear FL responses toward polyanions were designed. Their application to FL sensing of glycosaminoglycans (heparin: HEP, chondroitin 4-sulfate: ChS, and hyaluronic acid: HA) revealed that the charge density encoded as the unit structure directs the mode of OPV self-assembly: H-type aggregate for HEP with 16-times FL increase and J-type aggregate for HA with 93-times FL increase, thus unexpectedly achieving the preferential selectivity for HA in contrast to the conventional HEP selective systems. We have found that the integral magnitude of three factors consisting of binding mechanism, self-assembly, and FL response can amplify the structural information on the target input into the characteristic FL output. This emergent property has been used for a novel molecular recognition system that realizes unconventional FL sensing of HA, potentially applicable to the clinical diagnosis of cancer-related diseases. PMID:27060601

  8. Similarity recognition of molecular structures by optimal atomic matching and rotational superposition.

    PubMed

    Helmich, Benjamin; Sierka, Marek

    2012-01-15

    An algorithm for similarity recognition of molecules and molecular clusters is presented which also establishes the optimum matching among atoms of different structures. In the first step of the algorithm, a set of molecules are coarsely superimposed by transforming them into a common reference coordinate system. The optimum atomic matching among structures is then found with the help of the Hungarian algorithm. For this, pairs of structures are represented as complete bipartite graphs with a weight function that uses intermolecular atomic distances. In the final step, a rotational superposition method is applied using the optimum atomic matching found. This yields the minimum root mean square deviation of intermolecular atomic distances with respect to arbitrary rotation and translation of the molecules. Combined with an effective similarity prescreening method, our algorithm shows robustness and an effective quadratic scaling of computational time with the number of atoms. PMID:21997798

  9. A novel three-dimensional aerogel biochip for molecular recognition of nucleotide acids.

    PubMed

    Li, Yen Kuang; Yang, Den-Kai; Chen, Yun-Chu; Su, Hung-Ju; Wu, Jui-Chuang; Chen-Yang, Yui Whei

    2010-04-01

    Mesoporous aerogel was produced under regular atmospheric conditions using the sol-gel polymerization of tetraethyl orthosilicate with an ionic liquid as both solvent and active agent. This was then used to build a three-dimensional structure to recognize nucleotide acids. Fourier transformation infrared spectroscopy, scanning electron microscopy, (29)Si solid-state nuclear magnetic resonance, and Brunauer-Emmett-Teller instruments were used to characterize this 3D aerogel, demonstrating that it had high porosity and large internal networking surface area that could capture nucleotide acids. The functionality of molecular recognition on nucleotide acids was demonstrated by immobilizing an oligonucleotide to probe its DNA target and confirming the tagged fluorescent signals by confocal laser scanning microscopy. The results indicated that the as-prepared 3D bioaerogel was capable of providing a very large surface area to capture and recognize human gene ATP5O. PMID:19818421

  10. Molecular basis for oncohistone H3 recognition by SETD2 methyltransferase.

    PubMed

    Yang, Shuang; Zheng, Xiangdong; Lu, Chao; Li, Guo-Min; Allis, C David; Li, Haitao

    2016-07-15

    High-frequency point mutations of genes encoding histones have been identified recently as novel drivers in a number of tumors. Specifically, the H3K36M/I mutations were shown to be oncogenic in chondroblastomas and undifferentiated sarcomas by inhibiting H3K36 methyltransferases, including SETD2. Here we report the crystal structures of the SETD2 catalytic domain bound to H3K36M or H3K36I peptides with SAH (S-adenosylhomocysteine). In the complex structure, the catalytic domain adopts an open conformation, with the K36M/I peptide snuggly positioned in a newly formed substrate channel. Our structural and biochemical data reveal the molecular basis underying oncohistone recognition by and inhibition of SETD2. PMID:27474439

  11. Effect of Na+ binding on the conformation, stability and molecular recognition properties of thrombin

    PubMed Central

    De Filippis, Vincenzo; De Dea, Elisa; Lucatello, Filippo; Frasson, Roberta

    2005-01-01

    In the present work, the effect of Na+ binding on the conformational, stability and molecular recognition properties of thrombin was investigated. The binding of Na+ reduces the CD signal in the far-UV region, while increasing the intensity of the near-UV CD and fluorescence spectra. These spectroscopic changes have been assigned to perturbations in the environment of aromatic residues at the level of the S2 and S3 sites, as a result of global rigidification of the thrombin molecule. Indeed, the Na+-bound form is more stable to urea denaturation than the Na+-free form by ∼2 kcal/mol (1 cal≡4.184 J). Notably, the effects of cation binding on thrombin conformation and stability are specific to Na+ and parallel the affinity order of univalent cations for the enzyme. The Na+-bound form is even more resistant to limited proteolysis by subtilisin, at the level of the 148-loop, which is suggestive of the more rigid conformation this segment assumes in the ‘fast’ form. Finally, we have used hirudin fragment 1–47 as a molecular probe of the conformation of thrombin recognition sites in the fast and ‘slow’ form. From the effects of amino acid substitutions on the affinity of fragment 1–47 for the enzyme allosteric forms, we concluded that the specificity sites of thrombin in the Na+-bound form are in a more open and permissible conformation, compared with the more closed structure they assume in the slow form. Taken together, our results indicate that the binding of Na+ to thrombin serves to stabilize the enzyme into a more open and rigid conformation. PMID:15971999

  12. Engineering responsive polymer building blocks with host-guest molecular recognition for functional applications.

    PubMed

    Hu, Jinming; Liu, Shiyong

    2014-07-15

    CONSPECTUS: All living organisms and soft matter are intrinsically responsive and adaptive to external stimuli. Inspired by this fact, tremendous effort aiming to emulate subtle responsive features exhibited by nature has spurred the invention of a diverse range of responsive polymeric materials. Conventional stimuli-responsive polymers are constructed via covalent bonds and can undergo reversible or irreversible changes in chemical structures, physicochemical properties, or both in response to a variety of external stimuli. They have been imparted with a variety of emerging applications including drug and gene delivery, optical sensing and imaging, diagnostics and therapies, smart coatings and textiles, and tissue engineering. On the other hand, in comparison with molecular chemistry held by covalent bonds, supramolecular chemistry built on weak and reversible noncovalent interactions has emerged as a powerful and versatile strategy for materials fabrication due to its facile accessibility, extraordinary reversibility and adaptivity, and potent applications in diverse fields. Typically involving more than one type of noncovalent interactions (e.g., hydrogen bonding, metal coordination, hydrophobic association, electrostatic interactions, van der Waals forces, and π-π stacking), host-guest recognition refers to the formation of supramolecular inclusion complexes between two or more entities connected together in a highly controlled and cooperative manner. The inherently reversible and adaptive nature of host-guest molecular recognition chemistry, stemming from multiple noncovalent interactions, has opened up a new platform to construct novel types of stimuli-responsive materials. The introduction of host-guest chemistry not only enriches the realm of responsive materials but also confers them with promising new applications. Most intriguingly, the integration of responsive polymer building blocks with host-guest recognition motifs will endow the former with

  13. Tailoring molecularly imprinted polymer beads for alternariol recognition and analysis by a screening with mycotoxin surrogates.

    PubMed

    Abou-Hany, Rahma A G; Urraca, Javier L; Descalzo, Ana B; Gómez-Arribas, Lidia N; Moreno-Bondi, María C; Orellana, Guillermo

    2015-12-18

    Molecularly imprinted porous polymer microspheres have been prepared for selective binding of alternariol (AOH), a phenolic mycotoxin produced by Alternaria fungi. In order to lead the synthesis of recognition materials, four original AOH surrogates have been designed, prepared and characterized. They bear different number of phenol groups in various positions and different degree of O-methylation on the dibenzo[b,d]pyran-6-one skeleton. A comprehensive library of mixtures of basic, acidic or neutral monomers, with divinylbenzene or ethyleneglycol dimethacrylate as cross-linkers, were polymerized at a small scale in the presence of the four molecular mimics of the toxin molecule. This polymer screening has allowed selection of the optimal composition of the microbeads (N-(2-aminoethyl)methacrylamide, EAMA, and ethylene glycol dimethacrylate). The latter are able to bind AOH in water-acetonitrile (80:20, v/v) with an affinity constant of 109±10mM(-1) and a total number of binding sites of 35±2μmolg(-1), being alternariol monomethylether the only competitor species. Moreover, (1)H NMR titrations have unveiled a 1:2 surrogate-to-EAMA stoichiometry, the exact interaction sites and a binding constant of 1.5×10(4)M(-2). A molecularly imprinted solid phase extraction (MISPE) method has been optimized for selective isolation of the mycotoxin from aqueous samples upon a discriminating wash with 3mL of acetonitrile/water (20:80, v/v) followed by determination by HPLC with fluorescence detection. The method has been applied, in combination to ultrasound-assisted extraction, to the analysis of AOH in tomato samples fortified with the mycotoxin at five concentration levels (33-110μgkg(-1)), with recoveries in the range of 81-103% (RSD n=6). To the best of our knowledge, this is the first imprinted material capable of molecularly recognizing this widespread food contaminant. PMID:26632518

  14. Site-specific basicities regulate molecular recognition in receptor binding: in silico docking of thyroid hormones.

    PubMed

    Tóth, Gergő; Baska, Ferenc; Schretner, András; Rácz, Akos; Noszál, Béla

    2013-09-01

    Interactions between thyroid hormone α and β receptors and the eight protonation microspecies of each of the main thyroid hormones (thyroxine, liothyronine, and reverse liothyronine) were investigated and quantitated by molecular modeling. Flexible docking of the various protonation forms of thyroid hormones and high-affinity thyromimetics to the two thyroid receptors was carried out. In this method the role of the ionization state of each basic site could be studied in the composite process of molecular recognition. Our results quantitate at the molecular level how the ionization state and the charge distribution influence the protein binding. The anionic form of the carboxyl group (i.e., carboxylate site) is essential for protein binding, whereas the protonated form of amino group worsens the binding. The protonation state of the phenolate plays a less important role in the receptor affinity; its protonation, however, alters the electron density and the concomitant stacking propensity of the aromatic rings, resulting in a different binding score. The combined results of docking and microspeciation studies show that microspecies with the highest concentration at the pH of blood are not the strongest binding ones. The calculated binding free energy values can be well interpreted in terms of the interactions between the actual sites of the microspecies and the receptor amino acids. Our docking results were validated and compared with biological data from the literature. Since the thyroid hormone receptors influence several physiologic functions, such as metabolic rate, cholesterol and triglyceride levels, and heart frequency, our binding results provide a molecular basis for drug design and development in related therapeutic indications. PMID:23907234

  15. The role of protein "Stability patches" in molecular recognition: A case study of the human growth hormone-receptor complex.

    PubMed

    Osman, Roman; Mezei, Mihaly; Engel, Stanislav

    2016-04-15

    Dynamic characteristics of protein surfaces are among the factors determining their functional properties, including their potential participation in protein-protein interactions. The presence of clusters of static residues-"stability patches" (SPs)-is a characteristic of protein surfaces involved in intermolecular recognition. The mechanism, by with SPs facilitate molecular recognition, however, remains unclear. Analyzing the surface dynamic properties of the growth hormone and of its high-affinity variant we demonstrated that reshaping of the SPs landscape may be among the factors accountable for the improved affinity of this variant to the receptor. We hypothesized that SPs facilitate molecular recognition by moderating the conformational entropy of the unbound state, diminishing enthalpy-entropy compensation upon binding, and by augmenting the favorable entropy of desolvation. SPs mapping emerges as a valuable tool for investigating the structural basis of the stability of protein complexes and for rationalizing experimental approaches, such as affinity maturation, aimed at improving it. PMID:26691434

  16. Serine versus threonine glycosylation with α-O-GalNAc: unexpected selectivity in their molecular recognition with lectins.

    PubMed

    Madariaga, David; Martínez-Sáez, Nuria; Somovilla, Víctor J; García-García, Laura; Berbis, M Álvaro; Valero-Gónzalez, Jessika; Martín-Santamaría, Sonsoles; Hurtado-Guerrero, Ramon; Asensio, Juan L; Jiménez-Barbero, Jesús; Avenoza, Alberto; Busto, Jesús H; Corzana, Francisco; Peregrina, Jesús M

    2014-09-22

    The molecular recognition of several glycopeptides bearing Tn antigen (α-O-GalNAc-Ser or α-O-GalNAc-Thr) in their structure by three lectins with affinity for this determinant has been analysed. The work yields remarkable results in terms of epitope recognition, showing that the underlying amino acid of Tn (serine or threonine) plays a key role in the molecular recognition. In fact, while Soybean agglutinin and Vicia villosa agglutinin lectins prefer Tn-threonine, Helix pomatia agglutinin shows a higher affinity for the glycopeptides carrying Tn-serine. The different conformational behaviour of the two Tn biological entities, the residues of the studied glycopeptides in the close proximity to the Tn antigen and the topology of the binding site of the lectins are at the origin of these differences. PMID:25111627

  17. Linear Supramolecular Polymers via Connecting Telechelic Polycaprolactone through Alkynylplatinum(II) Terpyridine Molecular Tweezer/Pyrene Recognition Motif.

    PubMed

    Liu, Huaqing; Han, Xiaohang; Gao, Zongchun; Gao, Zhao; Wang, Feng

    2016-04-01

    By anchoring alkynylplatinum(II) terpyridine molecular tweezer/pyrene recognition motif on the chain-ends of telechelic polycaprolactone, high-molecular-weight supramolecular polymers have been successfully constructed via noncovalent chain extension, which demonstrate fascinating rheological and thermal properties. Moreover, the resulting assemblies exhibit interesting temperature- and solvent-responsive behaviors, which are promising for the development of adaptive functional materials. PMID:26924177

  18. Multibody cofactor and substrate molecular recognition in the myo-inositol monophosphatase enzyme.

    PubMed

    Ferruz, Noelia; Tresadern, Gary; Pineda-Lucena, Antonio; De Fabritiis, Gianni

    2016-01-01

    Molecular recognition is rarely a two-body protein-ligand problem, as it often involves the dynamic interplay of multiple molecules that together control the binding process. Myo-inositol monophosphatase (IMPase), a drug target for bipolar disorder, depends on 3 Mg(2+) ions as cofactor for its catalytic activity. Although the crystallographic pose of the pre-catalytic complex is well characterized, the binding process by which substrate, cofactor and protein cooperate is essentially unknown. Here, we have characterized cofactor and substrate cooperative binding by means of large-scale molecular dynamics. Our study showed the first and second Mg(2+) ions identify the binding pocket with fast kinetics whereas the third ion presents a much higher energy barrier. Substrate binding can occur in cooperation with cofactor, or alone to a binary or ternary cofactor-IMPase complex, although the last scenario occurs several orders of magnitude faster. Our atomic description of the three-body mechanism offers a particularly challenging example of pathway reconstruction, and may prove particularly useful in realistic contexts where water, ions, cofactors or other entities cooperate and modulate the binding process. PMID:27440438

  19. Multibody cofactor and substrate molecular recognition in the myo-inositol monophosphatase enzyme

    PubMed Central

    Ferruz, Noelia; Tresadern, Gary; Pineda-Lucena, Antonio; De Fabritiis, Gianni

    2016-01-01

    Molecular recognition is rarely a two-body protein-ligand problem, as it often involves the dynamic interplay of multiple molecules that together control the binding process. Myo-inositol monophosphatase (IMPase), a drug target for bipolar disorder, depends on 3 Mg2+ ions as cofactor for its catalytic activity. Although the crystallographic pose of the pre-catalytic complex is well characterized, the binding process by which substrate, cofactor and protein cooperate is essentially unknown. Here, we have characterized cofactor and substrate cooperative binding by means of large-scale molecular dynamics. Our study showed the first and second Mg2+ ions identify the binding pocket with fast kinetics whereas the third ion presents a much higher energy barrier. Substrate binding can occur in cooperation with cofactor, or alone to a binary or ternary cofactor-IMPase complex, although the last scenario occurs several orders of magnitude faster. Our atomic description of the three-body mechanism offers a particularly challenging example of pathway reconstruction, and may prove particularly useful in realistic contexts where water, ions, cofactors or other entities cooperate and modulate the binding process. PMID:27440438

  20. Electrochemical sensor for dopamine based on a novel graphene-molecular imprinted polymers composite recognition element.

    PubMed

    Mao, Yan; Bao, Yu; Gan, Shiyu; Li, Fenghua; Niu, Li

    2011-10-15

    A novel composite of graphene sheets/Congo red-molecular imprinted polymers (GSCR-MIPs) was synthesized through free radical polymerization (FRP) and applied as a molecular recognition element to construct dopamine (DA) electrochemical sensor. The template molecules (DA) were firstly absorbed at the GSCR surface due to their excellent affinity, and subsequently, selective copolymerization of methacrylic acid (MAA) and ethylene glycol dimethacrylate (EGDMA) was further achieved at the GSCR surface. Potential scanning was presented to extract DA molecules from the imprinted polymers film, and as a result, DA could be rapidly and completely removed by this way. With regard to the traditional MIPs, the GSCR-MIPs not only possessed a faster desorption and adsorption dynamics, but also exhibited a higher selectivity and binding capacity toward DA molecule. As a consequence, an electrochemical sensor for highly sensitive and selective detection of DA was successfully constructed as demonstration based on the synthesized GSCR-MIPs nanocomposites. Under experimental conditions, selective detection of DA in a linear concentration range of 1.0 × 10(-7)-8.3 × 10(-4)M was obtained, which revealed a lower limit of detection and wider linear response compared to some previously reported DA electrochemical MIPs sensors. The new DA electrochemical sensor based on GSCR-MIPs composites also exhibited excellent repeatability, which expressed as relative standard deviation (RSD) was about 2.50% for 30 repeated analyses of 20 μM DA. PMID:21824760

  1. Phthalocyanines as Molecular Scaffolds to Block Disease-Associated Protein Aggregation.

    PubMed

    Valiente-Gabioud, Ariel A; Miotto, Marco C; Chesta, María E; Lombardo, Verónica; Binolfi, Andres; Fernández, Claudio O

    2016-05-17

    amyloidogenic proteins. Analysis of the structure-activity relationship in phthalocyanines revealed that their anti-amyloid activity is highly dependent on the type of metal ion coordinated to the tetrapyrrolic system but is not sensitive to the number of peripheral charged substituents. The tendency of phthalocyanines to oligomerize (self-association) via aromatic-aromatic stacking interactions correlates precisely with their binding capabilities to target proteins and, more importantly, determines their efficiency as anti-amyloid agents. The ability to block different types of disease-associated protein aggregation raises the possibility that these cyclic tetrapyrrole compounds have a common mechanism of action to impair the formation of a variety of pathological aggregates. Because the structural and molecular basis for the anti-amyloid effects of these molecules is starting to emerge, combined efforts from the fields of structural, cellular, and animal biology will result critical for the rational design and discovery of new drugs for the treatment of amyloid related neurological disorders. PMID:27136297

  2. Molecularly imprinted polymer for recognition of 5-fluorouracil by RNA-type nucleobase pairing.

    PubMed

    Huynh, Tan-Phat; Pieta, Piotr; D'Souza, Francis; Kutner, Wlodzimierz

    2013-09-01

    A 6-aminopurine (adenine) derivative of bis(2,2'-bithienyl)methane, vis., 4-[2-(6-amino-9H-purin-9-yl)ethoxy]phenyl-4-[bis(2,2'-bithienyl)methane] or Ade-BTM, was designed and synthesized for recognition of 5-fluorouracil (FU), an antitumor chemotherapy agent, by RNA-type (nucleobase pairing)-driven molecular imprinting. The prepolymerization complex stoichiometry involved one FU molecule and two molecules of the Ade-BTM functional monomer. Molecular structure of this complex was thermodynamically optimized via density functional theory at the B3LYP/3-21G* level. The stability constant of the FU-Ade-BTM complex of 1:2 stoichiometry was K = 2.17(±0.07) × 10(7) M(-2), as determined by titration with quenching of fluorescence of the bis(2,2'-bithienyl)methane moiety of Ade-BTM by the FU titrant, in benzonitrile, at 352 nm excitation. Next, (5-fluorouracil)-templated molecularly imprinted polymer (MIP-FU) films were deposited on indium-tin oxide (ITO) or Au film-coated glass slides, Pt disk electrodes, or 10-MHz quartz crystal resonators by potentiodynamic electropolymerization from solution of FU, Ade-BTM, and tris([2,2'-bithiophen]-5-yl)methane (TTM) cross-linking monomer at FU:Ade-BTM:TTM = 1:2:3 mol ratio. Then UV-visible and Fourier transform infrared (FT-IR) spectra of the MIP-FU films were recorded to confirm the FU template presence in the MIP-FU film and its subsequent release by extraction with methanol from this film. For determination of the stability constant of the complex of the MIP cavity and FU, piezoelectric microgravimetry (PM) under both batch- and flow-injection analysis conditions was used. For sensing application, three different transduction platforms [differential pulse voltammetry (DPV), capacitive impedimetry (CI), and PM] were integrated with the MIP-FU recognition unit. The limit of detection (LOD) was 56 nM, 75 nM, and 0.26 mM, for these chemosensors, respectively, indicating suitability of the former two for FU determination in blood

  3. pH-responsive supramolecular polymerization in aqueous media driven by electrostatic attraction-enhanced crown ether-based molecular recognition.

    PubMed

    Ji, Xiaofan; Zhu, Kelong; Yan, Xuzhou; Ma, Yingjie; Li, Jinying; Hu, Bingjie; Yu, Yihua; Huang, Feihe

    2012-07-26

    All the previously reported supramolecular polymers based on crown ether-based molecular recognition have been prepared in anhydrous organic solvents. This is mainly due to the weakness of crown ether-based molecular recognition in the presence of water. Here we report a linear supramolecular polymer constructed from a heteroditopic monomer in an aqueous medium driven by crown ether-based molecular recognition through the introduction of electrostatic attraction. In addition, the reversible transition between the linear supramolecular polymer and oligomers is achieved by adding acid and base. This study realizes the breakthrough of the solvent for supramolecular polymerization driven by crown ether-based molecular recognition from anhydrous organic solvents to aqueous media. It is helpful for achieving supramolecular polymerization driven by crown ether-based molecular recognition in a completely aqueous medium. PMID:22495805

  4. Molecular recognition of genomic DNA in a condensate with a model surfactant for potential gene-delivery applications.

    PubMed

    Singh, Priya; Choudhury, Susobhan; Chandra, Goutam Kumar; Lemmens, Peter; Pal, Samir Kumar

    2016-04-01

    The functionality of a gene carrying nucleic acid in an artificial gene-delivery system is important for the overall efficiency of the vehicle in vivo. Here, we have studied a well-known artificial gene-delivery system, which is a condensate of calf thymus DNA (CT-DNA) with a model cationic surfactant cetyltrimethylammonium bromide (CTAB) to investigate the molecular recognition of the genomic DNA in the condensate. While dynamic light scattering (DLS) and circular dichroism (CD) reveal structural aspects of the condensate and the constituting DNA respectively, picosecond resolved polarization gated spectroscopy and Förster resonance energy transfer (FRET) reveal molecular recognition of the genomic DNA in the condensate. We have considered ethidium bromide (EB) and crystal violet (CV), which are well known DNA-binding agents through intercalative (specific) and electrostatic (non-specific) interactions, respectively, as model ligands for the molecular recognition studies. A fluorescent cationic surfactant, Nonyl Acridine Orange (NAO) is considered to be a mimic of CTAB in the condensate. The polarization gated fluorescence of NAO at various temperatures has been used to investigate the local microviscosity of the condensate. The excellent spectral overlap of NAO emission and the absorption spectra of both EB and CV allow us to investigate FRET-distances of the ligands with respect to NAO in the condensate at various temperatures and thermal stability of ligand-binding of the genomic DNA. The thermodynamic properties of the molecular recognition have also been explored using Van't Hoff equation. We have also extended our studies to molecular recognition of the genomic DNA in the condensate as dried thin films. This has important implications for its application in bioelectronics. PMID:26907719

  5. Designing and preparation of cytisine alkaloid surface-imprinted material and its molecular recognition characteristics

    NASA Astrophysics Data System (ADS)

    Gao, Baojiao; Bi, Concon; Fan, Li

    2015-03-01

    Based on molecular design, a cytisine surface-imprinted material was prepared using the new surface-imprinting technique of "pre-graft polymerizing and post-imprinting". The graft-polymerization of glycidyl methacrylate (GMA) on the surfaces of micron-sized silica gel particles was first performed with a surface-initiating system, preparing the grafted particles PGMA/SiO2. Subsequently, a polymer reaction, the ring-opening reaction of the epoxy groups of the grafted PGMA, was conducted with sodium 2,4-diaminobenzene sulfonate (SAS) as reagent, resulting in the functional grafted particles SAS-PGMA/SiO2. The adsorption of cytisine on SAS-PGMA/SiO2 particles reached saturation via strong electrostatic interaction between the sulfonate groups of SAS-PGMA/SiO2 particles and the protonated N atoms in cytisine molecule. Finally, cytisine surface-imprinting was successfully carried out with glutaraldehyde as crosslinker, obtaining cytisine surface-imprinted material MIP-SASP/SiO2. The binding and recognition characteristics of MIP-SASP/SiO2 towards cytisine were investigated in depth. The experimental results show that there is strong electrostatic interaction between particles and cytisine molecules, and on this basis, cytisine surface-imprinting can be smoothly performed. The surface-imprinted MIP-SASP/SiO2 has special recognition selectivity and excellent binding affinity for cytisine, and the selectivity coefficients of MIP-SASP/SiO2 particles for cytisine relative to matrine and oxymatrine, which were used as two contrast alkaloids, are 9.5 and 6.5, respectively.

  6. Molecular Dissection of Xyloglucan Recognition in a Prominent Human Gut Symbiont

    PubMed Central

    Tauzin, Alexandra S.; Kwiatkowski, Kurt J.; Orlovsky, Nicole I.; Smith, Christopher J.; Creagh, A. Louise; Haynes, Charles A.; Wawrzak, Zdzislaw

    2016-01-01

    ABSTRACT Polysaccharide utilization loci (PUL) within the genomes of resident human gut Bacteroidetes are central to the metabolism of the otherwise indigestible complex carbohydrates known as “dietary fiber.” However, functional characterization of PUL lags significantly behind sequencing efforts, which limits physiological understanding of the human-bacterial symbiosis. In particular, the molecular basis of complex polysaccharide recognition, an essential prerequisite to hydrolysis by cell surface glycosidases and subsequent metabolism, is generally poorly understood. Here, we present the biochemical, structural, and reverse genetic characterization of two unique cell surface glycan-binding proteins (SGBPs) encoded by a xyloglucan utilization locus (XyGUL) from Bacteroides ovatus, which are integral to growth on this key dietary vegetable polysaccharide. Biochemical analysis reveals that these outer membrane-anchored proteins are in fact exquisitely specific for the highly branched xyloglucan (XyG) polysaccharide. The crystal structure of SGBP-A, a SusD homolog, with a bound XyG tetradecasaccharide reveals an extended carbohydrate-binding platform that primarily relies on recognition of the β-glucan backbone. The unique, tetra-modular structure of SGBP-B is comprised of tandem Ig-like folds, with XyG binding mediated at the distal C-terminal domain. Despite displaying similar affinities for XyG, reverse-genetic analysis reveals that SGBP-B is only required for the efficient capture of smaller oligosaccharides, whereas the presence of SGBP-A is more critical than its carbohydrate-binding ability for growth on XyG. Together, these data demonstrate that SGBP-A and SGBP-B play complementary, specialized roles in carbohydrate capture by B. ovatus and elaborate a model of how vegetable xyloglucans are accessed by the Bacteroidetes. PMID:27118585

  7. Molecular recognition features (MoRFs) in three domains of life.

    PubMed

    Yan, Jing; Dunker, A Keith; Uversky, Vladimir N; Kurgan, Lukasz

    2016-03-01

    Intrinsically disordered proteins and protein regions offer numerous advantages in the context of protein-protein interactions when compared to the structured proteins and domains. These advantages include ability to interact with multiple partners, to fold into different conformations when bound to different partners, and to undergo disorder-to-order transitions concomitant with their functional activity. Molecular recognition features (MoRFs) are widespread elements located in disordered regions that undergo disorder-to-order transition upon binding to their protein partners. We characterize abundance, composition, and functions of MoRFs and their association with the disordered regions across 868 species spread across Eukaryota, Bacteria and Archaea. We found that although disorder is substantially elevated in Eukaryota, MoRFs have similar abundance and amino acid composition across the three domains of life. The abundance of MoRFs is highly correlated with the amount of intrinsic disorder in Bacteria and Archaea but only modestly correlated in Eukaryota. Proteins with MoRFs have significantly more disorder and MoRFs are present in many disordered regions, with Eukaryota having more MoRF-free disordered regions. MoRF-containing proteins are enriched in the ribosome, nucleus, nucleolus and microtubule and are involved in translation, protein transport, protein folding, and interactions with DNAs. Our insights into the nature and function of MoRFs enhance our understanding of the mechanisms underlying the disorder-to-order transition and protein-protein recognition and interactions. The fMoRFpred method that we used to annotate MoRFs is available at http://biomine.ece.ualberta.ca/fMoRFpred/. PMID:26651072

  8. A cell-based immunobiosensor with engineered molecular recognition--Part I: Design feasibility.

    PubMed

    Pizziconi, V B; Page, D L

    1997-01-01

    A novel bioelectronic sensor is described in which living immune cells are transformed into unique biotransducer couples by engineering their molecular recognition for preselected antigens of clinical interest. This 'hybrid' biosensor, constructed with mast cells interfaced to a microfabricated thermoelectric device with the use of biomolecular linkages, is capable of detecting antigens in real time by transducing minute heat changes arising from antigen-induced mast cell activation processes. The thermoelectric approach was selected based upon preliminary bioenergetic calculations which indicated that metabolic changes arising from mast cell antigen recognition result in a significant increase in exothermic heat relative to basal metabolic conditions. Experimental studies confirmed that mast cell activation and degranulation can be discriminated theramally from basal metabolic activity. Results obtained from microcalorimetry experiments using cultured mast cells (MC/9) mucosal-like mast cell line), and harvested mast cells (rat peritoneal mast cells) indicated that detectable increases in heat output (-3 +/- 0.5 pW/cell, mean peak output) immediately followed cell activation. The construction of a miniature hybrid immunobiosensor device was made possible by bioelectronic coupling achieved with the use of cellular adhesive proteins that immobilized non-adherent (MC/9) cells as well as adherent (RBL-2H3 rat basophilic leukemia) cells to the thermopile. Results from preliminary tests conducted on a hybrid biosensor prototype validated the design feasibility of a miniature, living cell immunodiagnostic biosensor. Such cell-based hybrid biosensor approaches may greatly extend the capability for selective, rapid, on-site, antigen detection for a wide range of clinically relevant antigens and offer new approaches to in vitro diagnostics. PMID:9178514

  9. New molecular scaffolds for the design of Mycobacterium tuberculosis type II dehydroquinase inhibitors identified using ligand and receptor based virtual screening.

    PubMed

    Kumar, Ashutosh; Siddiqi, Mohammad Imran; Miertus, Stanislav

    2010-04-01

    Using ligand and receptor based virtual screening approaches we have identified potential virtual screening hits targeting type II dehydroquinase from Mycobacterium tuberculosis, an effective and validated anti-mycobacterial target. Initially, we applied a virtual screening workflow based on a combination of 2D structural fingerprints, 3D pharmacophore and molecular docking to identify compounds that rigidly match specific aspects of ligand bioactive conformation. Subsequently, the resulting compounds were ranked and prioritized using receptor interaction fingerprint based scoring and quantitative structure activity relationship model developed using already known actives. The virtual screening hits prioritized belong to several classes of molecular scaffolds with several available substitution positions that could allow chemical modification to enhance binding affinity. Finally, identified hits may be useful to a medicinal chemist or combinatorial chemist to pick up the new molecular starting points for medicinal chemistry optimization for the design of novel type II dehydroquinase inhibitors. PMID:19816720

  10. Room temperature ionic liquid-mediated molecularly imprinted polymer monolith for the selective recognition of quinolones in pork samples.

    PubMed

    Sun, Xiangli; He, Jia; Cai, Guorui; Lin, Anqing; Zheng, Wenjie; Liu, Xuan; Chen, Langxing; He, Xiwen; Zhang, Yukui

    2010-12-01

    A novel molecularly imprinted polymer monolith was prepared by the room temperature ionic liquid-mediated in situ molecular imprinting technique, using norfloxacin (NOR) as the template, methacrylic acid as the functional monomer, ethylene glycol dimethacrylate as the cross-linker. The optimal synthesis conditions and recognition properties of NOR-imprinted monolithic column were investigated. The results indicated that the imprinted monoliths exhibited good ability of selective recognition against the template and its structural analog. Using the fabricated material as solid-phase extraction sorbent, a sample pre-treatment procedure of molecularly imprinted solid-phase extraction coupling with HPLC was developed for determination of trace quinolone residues in animal tissues samples. The recoveries ranging from 78.16 to 93.50% for eight quinolones antibiotics such as marbofloxacin, NOR, ciprofloxacin, danofloxacin, difloxacin, oxolinic acid, flumequine and enrofloxacin were obtained. PMID:21082676

  11. Molecular Recognition of Methyl α-d-Mannopyranoside by Antifreeze (Glyco)Proteins

    PubMed Central

    2015-01-01

    Antifreeze proteins and glycoproteins [AF(G)Ps] have been well-known for more than three decades for their ability to inhibit the growth and recrystallization of ice through binding to specific ice crystal faces, and they show remarkable structural compatibility with specific ice crystal faces. Here, we show that the crystal growth faces of methyl α-d-mannopyranoside (MDM), a representative pyranose sugar, also show noteworthy structural compatibility with the known periodicities of AF(G)Ps. We selected fish AFGPs (AFGP8, AFGP1–5), and a beetle AFP (DAFP1) with increasing antifreeze activity as potential additives for controlling MDM crystal growth. Similar to their effects on ice growth, the AF(G)Ps can inhibit MDM crystal growth and recrystallization, and more significantly, the effectiveness for the AF(G)Ps are well correlated with their antifreeze activity. MDM crystals grown in the presence of AF(G)Ps are smaller and have better defined shapes and are of higher quality as indicated by single crystal X-ray diffraction and polarized microscopy than control crystals, but no new polymorphs of MDM were identified by single crystal X-ray diffraction, solid-state NMR, and attenuated total reflectance infrared spectroscopy. The observed changes in the average sizes of the MDM crystals can be related to the changes in the number of the MDM nuclei in the presence of the AF(G)Ps. The critical free energy change differences of the MDM nucleation in the absence and presence of the additives were calculated. These values are close to those of the ice nucleation in the presence of AF(G)Ps suggesting similar interactions are involved in the molecular recognition of MDM by the AF(G)Ps. To our knowledge this is the first report where AF(G)Ps have been used to control crystal growth of carbohydrates and on AFGPs controlling non-ice-like crystals. Our finding suggests MDM might be a possible alternative to ice for studying the detailed mechanism of AF

  12. Molecular Dynamics of β-Hairpin Models of Epigenetic Recognition Motifs

    PubMed Central

    Zheng, Xiange; Wu, Chuanjie; Ponder, Jay W.; Marshall, Garland R.

    2012-01-01

    The conformations and stabilities of the β-hairpin model peptides of Waters1,2 have been experimentally characterized as a function of lysine ε-methylation. These models were developed to explore molecular recognition of known epigenetic recognition motifs. This system offered an opportunity to computationally examine the role of cation-π interactions, desolvation of the ε-methylated ammonium groups, and aromatic/aromatic interactions on the observed differences in NMR spectra. AMOEBA, a second-generation force field4, was chosen as it includes both multipole electrostatics and polarizability thought to be essential to accurately characterize such interactions. Independent parameterization of ε-methylated amines was required from which aqueous solvation free energies were estimated and shown to agree with literature values. Molecular dynamics simulations (100 ns) using the derived parameters with model peptides, such as Ac-R-W-V-W-V-N-G-Orn-K(Me)n -I-L-Q-NH2, where n = 0, 1, 2, or 3, were conducted in explicit solvent. Distances between the centers of the indole rings of the two-tryptophan residues, 2 and 4, and the ε-methylated ammonium group on Lys-9 as well as the distance between the N- and C-termini were monitored to estimate the strength and orientation of the cation-π and aromatic/aromatic interactions. In agreement with the experimental data, the stability of the β-hairpin increased significantly with lysine ε-methylation. The ability of MD simulations to reproduce the observed NOEs for the four peptides was further estimated for the monopole-based force fields, AMBER, CHARMM, and OPLSAA. AMOEBA correctly predicted over 80% of the observed NOEs for all four peptides, while the three-monopole force fields were 40–50% predictive in only two cases and approximately 10% in the other ten examples. Preliminary analysis suggests that the decreased cost of desolvation of the substituted ammonium group significantly compensated for the reduced cation

  13. Iptycene-derived crown ether hosts for molecular recognition and self-assembly.

    PubMed

    Han, Ying; Meng, Zheng; Ma, Ying-Xian; Chen, Chuan-Feng

    2014-07-15

    CONSPECTUS: Synthetic macrocyclic hosts have played key roles in the development of host-guest chemistry. Crown ethers are a class of macrocyclic molecules with unique flexible structures. They have served as the first generation of synthetic hosts, and researchers have extensively studied them in molecular recognition. However, the flexible structures of simple crown ethers and their relatively limited modes of complexation with guests have limited the further applications of these molecules. In recent years, researchers have moved toward fabricating interlocking molecules, supramolecular polymers, and other assemblies with specific structures and properties. Therefore, researchers have developed more complex crown ether-based macrocyclic hosts with multicavity structures and multicomplexation modes that provide more diverse and sophisticated host-guest systems. In this Account, we summarize our research on the synthesis and characterization of iptycene-derived crown ether hosts, their use as host molecules, and their applications in self-assembled complexes. Iptycenes including triptycenes and pentiptycenes are a class of aromatic compounds with unique rigid three-dimensional structures. As a result, they are promising building blocks for the synthesis of novel macrocyclic hosts and the construction of novel self-assembled complexes with specific structures and properties. During the last several years, we have designed and synthesized a new class of iptycene-derived crown ether hosts including macrotricyclic polyethers, molecular tweezer-like hosts, and tritopic tris(crown ether) hosts, which are all composed of rigid iptycene building blocks linked by flexible crown ether chains. We have examined the complexation behavior of these hosts with different types of organic guest molecules. Unlike with conventional crown ethers, the combination of iptycene moieties and crown ether chains provides the iptycene-derived crown ether hosts with complexation properties

  14. Molecular recognition of DNA by ligands: Roughness and complexity of the free energy profile

    NASA Astrophysics Data System (ADS)

    Zheng, Wenwei; Vargiu, Attilio Vittorio; Rohrdanz, Mary A.; Carloni, Paolo; Clementi, Cecilia

    2013-10-01

    Understanding the molecular mechanism by which probes and chemotherapeutic agents bind to nucleic acids is a fundamental issue in modern drug design. From a computational perspective, valuable insights are gained by the estimation of free energy landscapes as a function of some collective variables (CVs), which are associated with the molecular recognition event. Unfortunately the choice of CVs is highly non-trivial because of DNA's high flexibility and the presence of multiple association-dissociation events at different locations and/or sliding within the grooves. Here we have applied a modified version of Locally-Scaled Diffusion Map (LSDMap), a nonlinear dimensionality reduction technique for decoupling multiple-timescale dynamics in macromolecular systems, to a metadynamics-based free energy landscape calculated using a set of intuitive CVs. We investigated the binding of the organic drug anthramycin to a DNA 14-mer duplex. By performing an extensive set of metadynamics simulations, we observed sliding of anthramycin along the full-length DNA minor groove, as well as several detachments from multiple sites, including the one identified by X-ray crystallography. As in the case of equilibrium processes, the LSDMap analysis is able to extract the most relevant collective motions, which are associated with the slow processes within the system, i.e., ligand diffusion along the minor groove and dissociation from it. Thus, LSDMap in combination with metadynamics (and possibly every equivalent method) emerges as a powerful method to describe the energetics of ligand binding to DNA without resorting to intuitive ad hoc reaction coordinates.

  15. Conformational Melding Permits a Conserved Binding Geometry in TCR Recognition of Foreign and Self Molecular Mimics

    SciTech Connect

    Borbulevych, Oleg Y.; Piepenbrink, Kurt H.; Baker, Brian M.

    2012-03-16

    Molecular mimicry between foreign and self Ags is a mechanism of TCR cross-reactivity and is thought to contribute to the development of autoimmunity. The {alpha}{beta} TCR A6 recognizes the foreign Ag Tax from the human T cell leukemia virus-1 when presented by the class I MHC HLA-A2. In a possible link with the autoimmune disease human T cell leukemia virus-1-associated myelopathy/tropical spastic paraparesis, A6 also recognizes a self peptide from the neuronal protein HuD in the context of HLA-A2. We found in our study that the complexes of the HuD and Tax epitopes with HLA-A2 are close but imperfect structural mimics and that in contrast with other recent structures of TCRs with self Ags, A6 engages the HuD Ag with the same traditional binding mode used to engage Tax. Although peptide and MHC conformational changes are needed for recognition of HuD but not Tax and the difference of a single hydroxyl triggers an altered TCR loop conformation, TCR affinity toward HuD is still within the range believed to result in negative selection. Probing further, we found that the HuD-HLA-A2 complex is only weakly stable. Overall, these findings help clarify how molecular mimicry can drive self/nonself cross-reactivity and illustrate how low peptide-MHC stability can permit the survival of T cells expressing self-reactive TCRs that nonetheless bind with a traditional binding mode.

  16. Synthesis and computational investigation of molecularly imprinted nanospheres for selective recognition of alpha-tocopherol succinate

    PubMed Central

    Piacham, Theeraphon; Nantasenamat, Chanin; Isarankura-Na-Ayudhya, Chartchalerm; Prachayasittikul, Virapong

    2013-01-01

    Molecularly imprinted polymers (MIPs) are macromolecular matrices that can mimic the functional properties of antibodies, receptors and enzymes while possessing higher durability. As such, these polymers are interesting materials for applications in biomimetic sensor, drug synthesis, drug delivery and separation. In this study, we prepared MIPs and molecularly imprinted nanospheres (MINs) as receptors with specific recognition properties toward tocopherol succinate (TPS) in comparison to tocopherol (TP) and tocopherol nicotinate (TPN). MIPs were synthesized using methacrylic acid (MAA) as functional monomer, ethylene glycol dimethacrylate (EGDMA) as crosslinking agent and dichloromethane or acetronitrile as porogenic solvent under thermal-induced polymerization condition. Results indicated that imprinted polymers of TPS-MIP, TP-MIP and TPN-MIP all bound specifically to their template molecules at 2 folds greater than the non-imprinted polymers. The calculated binding capacity of all MIP was approximately 2 mg per gram of polymer when using the optimal rebinding solvent EtOH:H2O (3:2, v/v). Furthermore, the MINs toward TPS and TP were prepared by precipitation polymerization that yielded particles that are 200-400 nm in size. The binding capacities of MINs to their templates were greater than that of the non-imprinted nanospheres when using the optimal rebinding solvent EtOH:H2O (4:1, v/v). Computer simulation was performed to provide mechanistic insights on the binding modalities of template-monomer complexes. In conclusion, we had successful prepared MIPs and MINs for binding specifically to TP and TPS. Such MIPs and MINs have great potential for industrial and medical applications, particularly for the selective separation of TP and TPS. PMID:26622214

  17. Molecular insights into the recognition of N-terminal histone modifications by the BRPF1 bromodomain.

    PubMed

    Poplawski, Amanda; Hu, Kaifeng; Lee, Woonghee; Natesan, Senthil; Peng, Danni; Carlson, Samuel; Shi, Xiaobing; Balaz, Stefan; Markley, John L; Glass, Karen C

    2014-04-17

    The monocytic leukemic zinc finger (MOZ) histone acetyltransferase (HAT) acetylates free histones H3, H4, H2A, and H2B in vitro and is associated with up-regulation of gene transcription. The MOZ HAT functions as a quaternary complex with the bromodomain-PHD finger protein 1 (BRPF1), inhibitor of growth 5 (ING5), and hEaf6 subunits. BRPF1 links the MOZ catalytic subunit to the ING5 and hEaf6 subunits, thereby promoting MOZ HAT activity. Human BRPF1 contains multiple effector domains with known roles in gene transcription, as well as chromatin binding and remodeling. However, the biological function of the BRPF1 bromodomain remains unknown. Our findings reveal novel interactions of the BRPF1 bromodomain with multiple acetyllysine residues on the N-terminus of histones and show that it preferentially selects for H2AK5ac, H4K12ac, and H3K14ac. We used chemical shift perturbation data from NMR titration experiments to map the BRPF1 bromodomain ligand binding pocket and identified key residues responsible for coordination of the post-translationally modified histones. Extensive molecular dynamics simulations were used to generate structural models of bromodomain-histone ligand complexes, to analyze hydrogen bonding and other interactions, and to calculate the binding free energies. Our results outline the molecular mechanism driving binding specificity of the BRPF1 bromodomain for discrete acetyllysine residues on the N-terminal histone tails. Together, these data provide insights into how histone recognition by the bromodomain directs the biological function of BRPF1, ultimately targeting the MOZ HAT complex to chromatin substrates. PMID:24333487

  18. Molecular insights into the recognition of N-terminal histone modifications by the BRPF1 bromodomain

    PubMed Central

    Poplawski, Amanda; Hu, Kaifeng; Lee, Woonghee; Natesan, Senthil; Peng, Danni; Carlson, Samuel; Shi, Xiaobing; Balaz, Stefan; Markley, John L.; Glass, Karen C.

    2014-01-01

    The monocytic leukemic zinc-finger (MOZ) histone acetyltransferase (HAT) acetylates free histones H3, H4, H2A, and H2B in vitro and is associated with up-regulation of gene transcription. The MOZ HAT functions as a quaternary complex with the bromodomain-PHD finger protein 1 (BRPF1), inhibitor of growth 5 (ING5), and hEaf6 subunits. BRPF1 links the MOZ catalytic subunit to the ING5 and hEaf6 subunits, thereby promoting MOZ HAT activity. Human BRPF1 contains multiple effector domains with known roles in gene transcription, and chromatin binding and remodeling. However, the biological function of the BRPF1 bromodomain remains unknown. Our findings reveal novel interactions of the BRPF1 bromodomain with multiple acetyllysine residues on the N-terminus of histones, and show it preferentially selects for H2AK5ac, H4K12ac and H3K14ac. We used chemical shift perturbation data from NMR titration experiments to map the BRPF1 bromodomain ligand binding pocket and identified key residues responsible for coordination of the post-translationally modified histones. Extensive molecular dynamics simulations were used to generate structural models of bromodomain-histone ligand complexes, to analyze H-bonding and other interactions, and to calculate the binding free energies. Our results outline the molecular mechanism driving binding specificity of the BRPF1 bromodomain for discrete acetyllysine residues on the N-terminal histone tails. Together these data provide insights on how histone recognition by the bromodomain directs the biological function of BRPF1, ultimately targeting the MOZ HAT complex to chromatin substrates. PMID:24333487

  19. Molecular basis for specific recognition of bacterial ligands by NAIP/NLRC4 inflammasomes.

    PubMed

    Tenthorey, Jeannette L; Kofoed, Eric M; Daugherty, Matthew D; Malik, Harmit S; Vance, Russell E

    2014-04-10

    NLR (nucleotide-binding domain [NBD]- and leucine-rich repeat [LRR]-containing) proteins mediate innate immune sensing of pathogens in mammals and plants. How NLRs detect their cognate stimuli remains poorly understood. Here, we analyzed ligand recognition by NLR apoptosis inhibitory protein (NAIP) inflammasomes. Mice express multiple highly related NAIP paralogs that recognize distinct bacterial proteins. We analyzed a panel of 43 chimeric NAIPs, allowing us to map the NAIP domain responsible for specific ligand detection. Surprisingly, ligand specificity was mediated not by the LRR domain, but by an internal region encompassing several NBD-associated α-helical domains. Interestingly, we find that the ligand specificity domain has evolved under positive selection in both rodents and primates. We further show that ligand binding is required for the subsequent co-oligomerization of NAIPs with the downstream signaling adaptor NLR family, CARD-containing 4 (NLRC4). These data provide a molecular basis for how NLRs detect ligands and assemble into inflammasomes. PMID:24657167

  20. Molecular Recognition of Corticotropin releasing Factor by Its G protein-coupled Receptor CRFR1

    SciTech Connect

    Pioszak, Augen A.; Parker, Naomi R.; Suino-Powell, Kelly; Xu, H. Eric

    2009-01-15

    The bimolecular interaction between corticotropin-releasing factor (CRF), a neuropeptide, and its type 1 receptor (CRFR1), a class B G-protein-coupled receptor (GPCR), is crucial for activation of the hypothalamic-pituitary-adrenal axis in response to stress, and has been a target of intense drug design for the treatment of anxiety, depression, and related disorders. As a class B GPCR, CRFR1 contains an N-terminal extracellular domain (ECD) that provides the primary ligand binding determinants. Here we present three crystal structures of the human CRFR1 ECD, one in a ligand-free form and two in distinct CRF-bound states. The CRFR1 ECD adopts the alpha-beta-betaalpha fold observed for other class B GPCR ECDs, but the N-terminal alpha-helix is significantly shorter and does not contact CRF. CRF adopts a continuous alpha-helix that docks in a hydrophobic surface of the ECD that is distinct from the peptide-binding site of other class B GPCRs, thereby providing a basis for the specificity of ligand recognition between CRFR1 and other class B GPCRs. The binding of CRF is accompanied by clamp-like conformational changes of two loops of the receptor that anchor the CRF C terminus, including the C-terminal amide group. These structural studies provide a molecular framework for understanding peptide binding and specificity by the CRF receptors as well as a template for designing potent and selective CRFR1 antagonists for therapeutic applications.

  1. Leukemia-Associated Mutations in Nucleophosmin Alter Recognition by CRM1: Molecular Basis of Aberrant Transport

    PubMed Central

    Arregi, Igor; Falces, Jorge; Olazabal-Herrero, Anne; Alonso-Mariño, Marián; Taneva, Stefka G.; Rodríguez, José A.; Urbaneja, María A.; Bañuelos, Sonia

    2015-01-01

    Nucleophosmin (NPM) is a nucleocytoplasmic shuttling protein, normally enriched in nucleoli, that performs several activities related to cell growth. NPM mutations are characteristic of a subtype of acute myeloid leukemia (AML), where mutant NPM seems to play an oncogenic role. AML-associated NPM mutants exhibit altered subcellular traffic, being aberrantly located in the cytoplasm of leukoblasts. Exacerbated export of AML variants of NPM is mediated by the nuclear export receptor CRM1, and due, in part, to a mutationally acquired novel nuclear export signal (NES). To gain insight on the molecular basis of NPM transport in physiological and pathological conditions, we have evaluated the export efficiency of NPM in cells, and present new data indicating that, in normal conditions, wild type NPM is weakly exported by CRM1. On the other hand, we have found that AML-associated NPM mutants efficiently form complexes with CRM1HA (a mutant CRM1 with higher affinity for NESs), and we have quantitatively analyzed CRM1HA interaction with the NES motifs of these mutants, using fluorescence anisotropy and isothermal titration calorimetry. We have observed that the affinity of CRM1HA for these NESs is similar, which may help to explain the transport properties of the mutants. We also describe NPM recognition by the import machinery. Our combined cellular and biophysical studies shed further light on the determinants of NPM traffic, and how it is dramatically altered by AML-related mutations. PMID:26091065

  2. Synthesis of an oligonucleotide with a nicotinamide mononucleotide residue and its molecular recognition in DNA helices.

    PubMed

    Göckel, A; Richert, C

    2015-11-01

    Nicotinamide adenine dinucleotide (NAD) is a pivotal redox cofactor of primary metabolism. Its redox reactivity is based on the nicotinamide mononucleotide (NMN) moiety. We investigated whether NMN(+) can engage in pairing interactions, when incorporated into an oligonucleotide. Here we describe the incorporation of NMN(+) at the 3'-terminus of an oligodeoxynucleotide via a phosphoramidate coupling in solution. The stability of duplexes and triplexes with the NMN(+)-containing strand was measured in UV-melting curves. While the melting points of duplexes with different bases facing the nicotinamide were similar, triplex stabilities varied greatly between different base combinations, suggesting specific pairing. The most stable triplexes were found when a guanine and an adenine were facing the NMN(+) residue. Their triplex melting points were higher than those of the corresponding triplexes with a thymidine residue at the same position. These results show that NMN(+) residues can be recognized selectively in DNA helices and are thus compatible with the molecular recognition in nucleic acids. PMID:26371420

  3. Protein-Carbohydrate Interactions Studied by NMR: From Molecular Recognition to Drug Design

    PubMed Central

    Fernández-Alonso, María del Carmen; Díaz, Dolores; Berbis, Manuel Álvaro; Marcelo, Filipa; Cañada, Javier; Jiménez-Barbero, Jesús

    2012-01-01

    Diseases that result from infection are, in general, a consequence of specific interactions between a pathogenic organism and the cells. The study of host-pathogen interactions has provided insights for the design of drugs with therapeutic properties. One area that has proved to be promising for such studies is the constituted by carbohydrates which participate in biological processes of paramount importance. On the one hand, carbohydrates have shown to be information carriers with similar, if not higher, importance than traditionally considered carriers as amino acids and nucleic acids. On the other hand, the knowledge on molecular recognition of sugars by lectins and other carbohydrate-binding proteins has been employed for the development of new biomedical strategies. Biophysical techniques such as X-Ray crystallography and NMR spectroscopy lead currently the investigation on this field. In this review, a description of traditional and novel NMR methodologies employed in the study of sugar-protein interactions is briefly presented in combination with a palette of NMR-based studies related to biologically and/or pharmaceutically relevant applications. PMID:23305367

  4. Molecular basis for the recognition of methylated adenines in RNA by the eukaryotic YTH domain

    PubMed Central

    Luo, Shukun; Tong, Liang

    2014-01-01

    Methylation of the N6 position of selected internal adenines (m6A) in mRNAs and noncoding RNAs is widespread in eukaryotes, and the YTH domain in a collection of proteins recognizes this modification. We report the crystal structure of the splicing factor YT521-B homology (YTH) domain of Zygosaccharomyces rouxii MRB1 in complex with a heptaribonucleotide with an m6A residue in the center. The m6A modification is recognized by an aromatic cage, being sandwiched between a Trp and Tyr residue and with the methyl group pointed toward another Trp residue. Mutations of YTH domain residues in the RNA binding site can abolish the formation of the complex, confirming the structural observations. These residues are conserved in the human YTH proteins that also bind m6A RNA, suggesting a conserved mode of recognition. Overall, our structural and biochemical studies have defined the molecular basis for how the YTH domain functions as a reader of methylated adenines. PMID:25201973

  5. Molecular basis for the recognition of methylated adenines in RNA by the eukaryotic YTH domain.

    PubMed

    Luo, Shukun; Tong, Liang

    2014-09-23

    Methylation of the N6 position of selected internal adenines (m(6)A) in mRNAs and noncoding RNAs is widespread in eukaryotes, and the YTH domain in a collection of proteins recognizes this modification. We report the crystal structure of the splicing factor YT521-B homology (YTH) domain of Zygosaccharomyces rouxii MRB1 in complex with a heptaribonucleotide with an m(6)A residue in the center. The m(6)A modification is recognized by an aromatic cage, being sandwiched between a Trp and Tyr residue and with the methyl group pointed toward another Trp residue. Mutations of YTH domain residues in the RNA binding site can abolish the formation of the complex, confirming the structural observations. These residues are conserved in the human YTH proteins that also bind m(6)A RNA, suggesting a conserved mode of recognition. Overall, our structural and biochemical studies have defined the molecular basis for how the YTH domain functions as a reader of methylated adenines. PMID:25201973

  6. A metal–ion-responsive adhesive material via switching of molecular recognition properties

    PubMed Central

    Nakamura, Takashi; Takashima, Yoshinori; Hashidzume, Akihito; Yamaguchi, Hiroyasu; Harada, Akira

    2014-01-01

    Common adhesives stick to a wide range of materials immediately after they are applied to the surfaces. To prevent indiscriminate sticking, smart adhesive materials that adhere to a specific target surface only under particular conditions are desired. Here we report a polymer hydrogel modified with both β-cyclodextrin (βCD) and 2,2′-bipyridyl (bpy) moieties (βCD–bpy gel) as a functional adhesive material responding to metal ions as chemical stimuli. The adhesive property of βCD–bpy gel based on interfacial molecular recognition is expressed by complexation of metal ions to bpy that controlled dissociation of supramolecular cross-linking of βCD–bpy. Moreover, adhesion of βCD–bpy gel exhibits selectivity on the kinds of metal ions, depending on the efficiency of metal–bpy complexes in cross-linking. Transduction of two independent chemical signals (metal ions and host–guest interactions) is achieved in this adhesion system, which leads to the development of highly orthogonal macroscopic joining of multiple objects. PMID:25099995

  7. In Vitro Selection of a Single-Stranded DNA Molecular Recognition Element against Atrazine

    PubMed Central

    Williams, Ryan M.; Crihfield, Cassandra L.; Gattu, Srikanth; Holland, Lisa A.; Sooter, Letha J.

    2014-01-01

    Widespread use of the chlorotriazine herbicide, atrazine, has led to serious environmental and human health consequences. Current methods of detecting atrazine contamination are neither rapid nor cost-effective. In this work, atrazine-specific single-stranded DNA (ssDNA) molecular recognition elements (MRE) were isolated. We utilized a stringent Systematic Evolution of Ligands by Exponential Enrichment (SELEX) methodology that placed the greatest emphasis on what the MRE should not bind to. After twelve rounds of SELEX, an atrazine-specific MRE with high affinity was obtained. The equilibrium dissociation constant (Kd) of the ssDNA sequence is 0.62 ± 0.21 nM. It also has significant selectivity for atrazine over atrazine metabolites and other pesticides found in environmentally similar locations and concentrations. Furthermore, we have detected environmentally relevant atrazine concentrations in river water using this MRE. The strong affinity and selectivity of the selected atrazine-specific ssDNA validated the stringent SELEX methodology and identified a MRE that will be useful for rapid atrazine detection in environmental samples. PMID:25196435

  8. Mapping molecular adhesion sites inside SMIL coated capillaries using atomic force microscopy recognition imaging.

    PubMed

    Leitner, Michael; Stock, Lorenz G; Traxler, Lukas; Leclercq, Laurent; Bonazza, Klaus; Friedbacher, Gernot; Cottet, Hervé; Stutz, Hanno; Ebner, Andreas

    2016-08-01

    Capillary zone electrophoresis (CZE) is a powerful analytical technique for fast and efficient separation of different analytes ranging from small inorganic ions to large proteins. However electrophoretic resolution significantly depends on the coating of the inner capillary surface. High technical efforts like Successive Multiple Ionic Polymer Layer (SMIL) generation have been taken to develop stable coatings with switchable surface charges fulfilling the requirements needed for optimal separation. Although the performance can be easily proven in normalized test runs, characterization of the coating itself remains challenging. Atomic force microscopy (AFM) allows for topographical investigation of biological and analytical relevant surfaces with nanometer resolution and yields information about the surface roughness and homogeneity. Upgrading the scanning tip to a molecular biosensor by adhesive molecules (like partly inverted charged molecules) allows for performing topography and recognition imaging (TREC). As a result, simultaneously acquired sample topography and adhesion maps can be recorded. We optimized this technique for electrophoresis capillaries and investigated the charge distribution of differently composed and treated SMIL coatings. By using the positively charged protein avidin as a single molecule sensor, we compared these SMIL coatings with respect to negative charges, resulting in adhesion maps with nanometer resolution. The capability of TREC as a functional investigation technique at the nanoscale was successfully demonstrated. PMID:27265903

  9. Atomic interactions of neonicotinoid agonists with AChBP: Molecular recognition of the distinctive electronegative pharmacophore

    SciTech Connect

    Talley, Todd T.; Harel, Michal; Hibbs, Ryan E.; Radi, Zoran; Tomizawa, Motohiro; Casida, John E.; Taylor, Palmer

    2008-07-28

    Acetylcholine-binding proteins (AChBPs) from mollusks are suitable structural and functional surrogates of the nicotinic acetylcholine receptors when combined with transmembrane spans of the nicotinic receptor. These proteins assemble as a pentamer with identical ACh binding sites at the subunit interfaces and show ligand specificities resembling those of the nicotinic receptor for agonists and antagonists. A subset of ligands, termed the neonicotinoids, exhibit specificity for insect nicotinic receptors and selective toxicity as insecticides. AChBPs are of neither mammalian nor insect origin and exhibit a distinctive pattern of selectivity for the neonicotinoid ligands. We define here the binding orientation and determinants of differential molecular recognition for the neonicotinoids and classical nicotinoids by estimates of kinetic and equilibrium binding parameters and crystallographic analysis. Neonicotinoid complex formation is rapid and accompanied by quenching of the AChBP tryptophan fluorescence. Comparisons of the neonicotinoids imidacloprid and thiacloprid in the binding site from Aplysia californica AChBP at 2.48 and 1.94 {angstrom} in resolution reveal a single conformation of the bound ligands with four of the five sites occupied in the pentameric crystal structure. The neonicotinoid electronegative pharmacophore is nestled in an inverted direction compared with the nicotinoid cationic functionality at the subunit interfacial binding pocket. Characteristic of several agonists, loop C largely envelops the ligand, positioning aromatic side chains to interact optimally with conjugated and hydrophobic regions of the neonicotinoid. This template defines the association of interacting amino acids and their energetic contributions to the distinctive interactions of neonicotinoids.

  10. Molecular recognition and controlled release in drug delivery systems based on nanostructured lipid surfactants

    NASA Astrophysics Data System (ADS)

    Angius, R.; Murgia, S.; Berti, D.; Baglioni, P.; Monduzzi, M.

    2006-08-01

    Several monoolein/water (MO/W) based liquid crystalline (LC) nanostructured mesophases have been revisited in view of the new trends of modern drug delivery formulations. The shape and amphiphilic character of the investigated lipid molecules address the preferential polar-apolar interfacial curvature and the delicate interplay of different intermolecular forces that drive self-assembly and thermodynamic stability of the nanostructures. Here some preliminary results related to the release of the antiviral drug 1-amine-adamantane hydrochloride, solubilized in the aqueous domain of bicontinuous cubic and reverse hexagonal LC phases, suggest these MO based LC phases as possible nano-depot systems for long term controlled release. Drug release was followed by conductivity measurements during a period of ten days. An effective and targeted drug delivery often requires a specific molecular recognition. With this aim, the possibility to entrap suitable molecules such as lauroylcholine (LCh, a cationic surfactant having a peptide-like polar head that can 'recognize' membrane proteins) and adenosine monophosphate disodium salt (NaAMP, an electrolyte that can 'recognize' purine receptors) has been tested. The addition of LCh to MO/W cubic gyroid (CG) LC phase causes a cubic-lamellar phase transition. The addition of NaAMP still allows the formation of the CG nanostructure. In the presence of both NaAMP and LCh again a CG LC phase forms. The bicontinuous CG LC phases have been characterized by NMR and SAXS.

  11. Molecular Mechanisms of Taste Recognition: Considerations about the Role of Saliva

    PubMed Central

    Fábián, Tibor Károly; Beck, Anita; Fejérdy, Pál; Hermann, Péter; Fábián, Gábor

    2015-01-01

    The gustatory system plays a critical role in determining food preferences and food intake, in addition to nutritive, energy and electrolyte balance. Fine tuning of the gustatory system is also crucial in this respect. The exact mechanisms that fine tune taste sensitivity are as of yet poorly defined, but it is clear that various effects of saliva on taste recognition are also involved. Specifically those metabolic polypeptides present in the saliva that were classically considered to be gut and appetite hormones (i.e., leptin, ghrelin, insulin, neuropeptide Y, peptide YY) were considered to play a pivotal role. Besides these, data clearly indicate the major role of several other salivary proteins, such as salivary carbonic anhydrase (gustin), proline-rich proteins, cystatins, alpha-amylases, histatins, salivary albumin and mucins. Other proteins like glucagon-like peptide-1, salivary immunoglobulin-A, zinc-α-2-glycoprotein, salivary lactoperoxidase, salivary prolactin-inducible protein and salivary molecular chaperone HSP70/HSPAs were also expected to play an important role. Furthermore, factors including salivary flow rate, buffer capacity and ionic composition of saliva should also be considered. In this paper, the current state of research related to the above and the overall emerging field of taste-related salivary research alongside basic principles of taste perception is reviewed. PMID:25782158

  12. Recombinase-based isothermal amplification of nucleic acids with self-avoiding molecular recognition systems (SAMRS).

    PubMed

    Sharma, Nidhi; Hoshika, Shuichi; Hutter, Daniel; Bradley, Kevin M; Benner, Steven A

    2014-10-13

    Recombinase polymerase amplification (RPA) is an isothermal method to amplify nucleic acid sequences without the temperature cycling that classical PCR uses. Instead of using heat to denature the DNA duplex, RPA uses recombination enzymes to swap single-stranded primers into the duplex DNA product; these are then extended using a strand-displacing polymerase to complete the cycle. Because RPA runs at low temperatures, it never forces the system to recreate base-pairs following Watson-Crick rules, and therefore it produces undesired products that impede the amplification of the desired product, complicating downstream analysis. Herein, we show that most of these undesired side products can be avoided if the primers contain components of a self-avoiding molecular recognition system (SAMRS). Given the precision that is necessary in the recombination systems for them to function biologically, it is surprising that they accept SAMRS. SAMRS-RPA is expected to be a powerful tool within the range of amplification techniques available to scientists. PMID:25209570

  13. Boronate affinity materials for separation and molecular recognition: structure, properties and applications.

    PubMed

    Li, Daojin; Chen, Yang; Liu, Zhen

    2015-11-21

    Boronate affinity materials, as unique sorbents, have emerged as important media for the selective separation and molecular recognition of cis-diol-containing compounds. With the introduction of boronic acid functionality, boronate affinity materials exhibit several significant advantages, including broad-spectrum selectivity, reversible covalent binding, pH-controlled capture/release, fast association/desorption kinetics, and good compatibility with mass spectrometry. Because cis-diol-containing biomolecules, including nucleosides, saccharides, glycans, glycoproteins and so on, are the important targets in current research frontiers such as metabolomics, glycomics and proteomics, boronate affinity materials have gained rapid development and found increasing applications in the last decade. In this review, we critically survey recent advances in boronate affinity materials. We focus on fundamental considerations as well as important progress and new boronate affinity materials reported in the last decade. We particularly discuss on the effects of the structure of boronate ligands and supporting materials on the properties of boronate affinity materials, such as binding pH, affinity, selectivity, binding capacity, tolerance for interference and so on. A variety of promising applications, including affinity separation, proteomics, metabolomics, disease diagnostics and aptamer selection, are introduced with main emphasis on how boronate affinity materials can solve the issues in the applications and what merits boronate affinity materials can provide. PMID:26377373

  14. Molecular basis of mycobacterial lipid antigen presentation by CD1c and its recognition by αβ T cells.

    PubMed

    Roy, Sobhan; Ly, Dalam; Li, Nan-Sheng; Altman, John D; Piccirilli, Joseph A; Moody, D Branch; Adams, Erin J

    2014-10-28

    CD1c is a member of the group 1 CD1 family of proteins that are specialized for lipid antigen presentation. Despite high cell surface expression of CD1c on key antigen-presenting cells and the discovery of its mycobacterial lipid antigen presentation capability, the molecular basis of CD1c recognition by T cells is unknown. Here we present a comprehensive functional and molecular analysis of αβ T-cell receptor (TCR) recognition of CD1c presenting mycobacterial phosphomycoketide antigens. Our structure of CD1c with the mycobacterial phosphomycoketide (PM) shows similarities to that of CD1c-mannosyl-β1-phosphomycoketide in that the A' pocket accommodates the mycoketide alkyl chain; however, the phosphate head-group of PM is shifted ∼6 Å in relation to that of mannosyl-β1-PM. We also demonstrate a bona fide interaction between six human TCRs and CD1c-mycoketide complexes, measuring high to moderate affinities. The crystal structure of the DN6 TCR and mutagenic studies reveal a requirement of five complementarity determining region (CDR) loops for CD1c recognition. Furthermore, mutagenesis of CD1c reveals residues in both the α1 and α2 helices involved in TCR recognition, yet not entirely overlapping among the examined TCRs. Unlike patterns for MHC I, no archetypical binding footprint is predicted to be shared by CD1c-reactive TCRs, even when recognizing the same or similar antigens. PMID:25298532

  15. Molecular basis of mycobacterial lipid antigen presentation by CD1c and its recognition by αβ T cells

    PubMed Central

    Roy, Sobhan; Ly, Dalam; Li, Nan-Sheng; Altman, John D.; Piccirilli, Joseph A.; Moody, D. Branch; Adams, Erin J.

    2014-01-01

    CD1c is a member of the group 1 CD1 family of proteins that are specialized for lipid antigen presentation. Despite high cell surface expression of CD1c on key antigen-presenting cells and the discovery of its mycobacterial lipid antigen presentation capability, the molecular basis of CD1c recognition by T cells is unknown. Here we present a comprehensive functional and molecular analysis of αβ T-cell receptor (TCR) recognition of CD1c presenting mycobacterial phosphomycoketide antigens. Our structure of CD1c with the mycobacterial phosphomycoketide (PM) shows similarities to that of CD1c-mannosyl-β1-phosphomycoketide in that the A' pocket accommodates the mycoketide alkyl chain; however, the phosphate head-group of PM is shifted ∼6 Å in relation to that of mannosyl-β1-PM. We also demonstrate a bona fide interaction between six human TCRs and CD1c-mycoketide complexes, measuring high to moderate affinities. The crystal structure of the DN6 TCR and mutagenic studies reveal a requirement of five complementarity determining region (CDR) loops for CD1c recognition. Furthermore, mutagenesis of CD1c reveals residues in both the α1 and α2 helices involved in TCR recognition, yet not entirely overlapping among the examined TCRs. Unlike patterns for MHC I, no archetypical binding footprint is predicted to be shared by CD1c-reactive TCRs, even when recognizing the same or similar antigens. PMID:25298532

  16. Inverse opal spheres based on polyionic liquids as functional microspheres with tunable optical properties and molecular recognition capabilities.

    PubMed

    Cui, Jiecheng; Zhu, Wei; Gao, Ning; Li, Jian; Yang, Haowei; Jiang, Yin; Seidel, Philipp; Ravoo, Bart Jan; Li, Guangtao

    2014-04-01

    Based on the combination of the unique features of both polyionic liquids and spherical colloidal crystals, a new class of inverse opaline spheres with a series of distinct properties was fabricated. It was found that such photonic spheres could not only be used as stimuli-responsive photonic microgels, but also serve as multifunctional microspheres that mimic the main characteristics of conventional molecules, including intrinsic optical properties, specific molecular recognition, reactivity and derivatization, and anisotropy. PMID:24596228

  17. Single molecular recognition force spectroscopy study of a DNA aptamer with the target epithelial cell adhesion molecule.

    PubMed

    Wang, Nan; Liu, Huiqing; Hao, Jinhui; Bai, Xiaojing; Li, Huiyan; Zhang, Zhe; Wang, Hongda; Tang, Jilin

    2015-09-21

    The epithelial cell adhesion molecule (EpCAM) is a tumor-specific antigen for malignancies of the epithelialis lineage. In this study the interaction between the DNA-based EpCAM aptamer (SYL3C) and EpCAM was explored using single molecular recognition force spectroscopy (SMFS). The capability of aptamer SYL3C to recognize the EpCAM protein and the kinetic parameters were investigated. PMID:26229987

  18. High-Throughput Screen in Cryptococcus neoformans Identifies a Novel Molecular Scaffold That Inhibits Cell Wall Integrity Pathway Signaling

    PubMed Central

    2015-01-01

    Cryptococcus neoformans is one of the most important human fungal pathogens; however, no new therapies have been developed in over 50 years. Fungicidal activity is crucially important for an effective anticryptococal agent and, therefore, we screened 361,675 molecules against C. neoformans using an adenylate kinase release assay that specifically detects fungicidal activity. A set of secondary assays narrowed the set of hits to molecules that interfere with fungal cell wall integrity and identified three benzothioureas with low in vitro mammalian toxicity and good in vitro anticryptococcal (minimum inhibitory concentration = 4 μg/mL). This scaffold inhibits signaling through the cell wall integrity MAP kinase cascade. Structure–activity studies indicate that the thiocarbonyl moiety is crucial for activity. Genetic and biochemical data suggest that benzothioureas inhibit signaling upstream of the kinase cascade. Thus, the benzothioureas appear to be a promising new scaffold for further exploration in the search for new anticryptococcal agents. PMID:26807437

  19. A core-shell surface magnetic molecularly imprinted polymers with fluorescence for λ-cyhalothrin selective recognition.

    PubMed

    Gao, Lin; Wang, Jixiang; Li, Xiuying; Yan, Yongsheng; Li, Chunxiang; Pan, Jianming

    2014-11-01

    In this study, we report here a general protocol for making core-shell magnetic Fe3O4/SiO2-MPS/MIPs (MPS = 3-(methacryloxyl) propyl trimethoxysilane, MIPs = molecularly imprinted polymers, Fe3O4/SiO2-MPS as core, MIPs as shell) via a surface molecular imprinting technique for optical detection of trace λ-cyhalothrin. The fluorescent molecularly imprinted polymer shell was first prepared by copolymerization of acrylamide with a small quantity of allyl fluorescein in the presence of λ-cyhalothrin to form recognition sites without doping. The magnetic Fe3O4/SiO2-MPS/MIPs exhibited paramagnetism, high fluorescence intensity, and highly selective recognition. Using fluorescence quenching as a detecting tool, Fe3O4/SiO2-MPS/MIPs were successfully applied to selectively and sensitively detect λ-cyhalothrin, and a linear relationship could be obtained covering a wide concentration range of 0-50 nM with a correlation coefficient of 0.9962 described by the Stern-Volmer equation. The experimental results of practical detection revealed that magnetic Fe3O4/SiO2-MPS/MIPs as an attractive recognition element was satisfactory for determination of trace λ-cyhalothrin in honey samples. This study, therefore, demonstrated the potential of MIPs for detection of λ-cyhalothrin in food. PMID:25200071

  20. Scaffolding and Metacognition

    ERIC Educational Resources Information Center

    Holton, Derek; Clarke, David

    2006-01-01

    This paper proposes an expanded conception of scaffolding with four key elements: (1) scaffolding agency--expert, reciprocal, and self-scaffolding; (2) scaffolding domain--conceptual and heuristic scaffolding; (3) the identification of self-scaffolding with metacognition; and (4) the identification of six zones of scaffolding activity; each zone…

  1. A promiscuous recognition mechanism between GPR17 and SDF-1: Molecular insights.

    PubMed

    Parravicini, Chiara; Daniele, Simona; Palazzolo, Luca; Trincavelli, Maria Letizia; Martini, Claudia; Zaratin, Paola; Primi, Roberto; Coppolino, Giusy; Gianazza, Elisabetta; Abbracchio, Maria P; Eberini, Ivano

    2016-06-01

    Recent data and publications suggest a promiscuous behaviour for GPR17, a class-A GPCR operated by different classes of ligands, such as uracil nucleotides, cysteinyl-leukotrienes and oxysterols. This observation, together with the ability of several class-A GPCRs to form homo- and hetero-dimers, is likely to unveil new pathophysiological roles and novel emerging pharmacological properties for some of these GPCRs, including GPR17. This receptor shares structural, phylogenetic and functional properties with some chemokine receptors, CXCRs. Both GPR17 and CXCR2 are operated by oxysterols, and both GPR17 and CXCR ligands have been demonstrated to have a role in orchestrating inflammatory responses and oligodendrocyte precursor cell differentiation to myelinating cells in acute and chronic diseases of the central nervous system. Here, by combining in silico modelling data with in vitro validation in (i) a classical reference pharmacological assay for GPCR activity and (ii) a model of maturation of primary oligodendrocyte precursor cells, we demonstrate that GPR17 can be activated by SDF-1, a ligand of chemokine receptors CXCR4 and CXCR7, and investigate the underlying molecular recognition mechanism. We also demonstrate that cangrelor, a GPR17 orthosteric antagonist, can block the SDF-1-mediated activation of GPR17 in a concentration-dependent manner. The ability of GPR17 to respond to different classes of GPCR ligands suggests that this receptor modifies its function depending on the extracellular mileu changes occurring under specific pathophysiological conditions and advocates it as a strategic target for neurodegenerative diseases with an inflammatory/immune component. PMID:26971834

  2. Molecular structure and peptidoglycan recognition of Mycobacterium tuberculosis ArfA (Rv0899)

    PubMed Central

    Yao, Yong; Barghava, Neha; Kim, Johnny; Niederweis, Michael; Marassi, Francesca M.

    2012-01-01

    M. tuberculosis ArfA (Rv0899) is a membrane protein encoded by an operon that is required for supporting bacterial growth in acidic environments. Its C-terminal domain (C domain) shares significant sequence homology with the OmpA-like family of peptidoglycan-binding domains, suggesting that its physiological function in acid stress protection may be related to its interaction with the mycobacterial cell wall. Previously, we showed that ArfA forms three independently structured modules and we reported the structure of its central domain (B domain). Here we describe the high-resolution structure and dynamics of the C domain, we identify ArfA as a peptidoglycan-binding protein, and elucidate the molecular basis for its specific recognition of diaminopimelate (DAP) type peptidoglycan. The C domain of ArfA adopts the characteristic fold of the OmpA-like family. It exhibits pH-dependent conformational dynamics (with significant hereogeneity at neutral pH and a more ordered structure at acidic pH), which could be related to its acid-stress response. The C domain associates tightly with polymeric peptidoglycan isolated from M. tuberculosis and also associates with a soluble peptide intermediate of peptidoglycan biosynthesis. This enabled us to characterize the peptidoglycan binding site where five highly conserved ArfA residues, including two key arginines, establish the specificity for DAP- but not Lys-type peptidoglycan. ArfA is the first peptidoglycan-binding protein to be identified in M. tuberculosis. Its functions in acid stress protection and peptidoglycan binding suggest a link between the acid stress response and the physico-chemical properties of the mycobacterial cell wall. PMID:22206986

  3. Probability model for molecular recognition in biological receptor repertoires: significance to the olfactory system.

    PubMed

    Lancet, D; Sadovsky, E; Seidemann, E

    1993-04-15

    A generalized phenomenological model is presented for stereospecific recognition between biological receptors and their ligands. We ask what is the distribution of binding constants psi(K) between an arbitrary ligand and members of a large receptor repertoire, such as immunoglobulins or olfactory receptors. For binding surfaces with B potential subsite and S different types of subsite configurations, the number of successful elementary interactions obeys a binomial distribution. The discrete probability function psi(K) is then derived with assumptions on alpha, the free energy contribution per elementary interaction. The functional form of psi(K) may be universal, although the parameter values could vary for different ligand types. An estimate of the parameter values of psi(K) for iodovanillin, an analog of odorants and immunological haptens, is obtained by equilibrium dialysis experiments with nonimmune antibodies. Based on a simple relationship, predicted by the model, between the size of a receptor repertoire and its average maximal affinity toward an arbitrary ligand, the size of the olfactory receptor repertoire (Nolf) is calculated as 300-1000, in very good agreement with recent molecular biological studies. A very similar estimate, Nolf = 500, is independently derived by relating a theoretical distribution of maxima for psi(K) with published human olfactory threshold variations. The present model also has implications to the question of olfactory coding and to the analysis of specific anosmias, genetic deficits in perceiving particular odorants. More generally, the proposed model provides a better understanding of ligand specificity in biological receptors and could help in understanding their evolution. PMID:8475121

  4. Biopolymeric receptor for peptide recognition by molecular imprinting approach--synthesis, characterization and application.

    PubMed

    Singh, Lav Kumar; Singh, Monika; Singh, Meenakshi

    2014-12-01

    The present work is focused on the development of a biocompatible zwitterionic hydrogel for various applications in analytical chemistry. Biopolymer chitosan was derivatized to obtain a series of zwitterionic hydrogel samples. Free amino groups hanging on the biopolymeric chain were reacted with γ-butyrolactone to quaternize the N-centers of polymeric chain. N,N-methylene-bis-acrylamide acts as a crosslinker via Michael-type addition in the subsequent step and facilitated gelation of betainized chitosan. These biopolymeric hydrogel samples were fully characterized by FTIR, (1)H NMR, (13)C NMR spectra, SEM and XRD. Hydrogels were further characterized for their swelling behavior at varying parameters. The extent of swelling was perceived to be dictated by solvent composition such as pH, ionic strength and temperature. This valuable polymeric format is herein chosen to design an artificial receptor for dipeptide 'carnosine', which has adequate societal significance to be analytically determined, by molecular imprinting. Electrostatic interactions along with complementary H-bonding and other hydrophobic interactions inducing additional synergetic effect between the template (carnosine) and the imprinted polymer led to the formation of imprinted sites. The MIP was able to selectively and specifically take up carnosine from aqueous solution quantitatively. Thus prepared MIPs were characterized by FTIR spectroscopy, SEM providing evidence for the quality and quantity of imprinted gels. The binding studies showed that the MIP illustrated good recognition for carnosine as compared to non-imprinted polymers (NIPs). Detection limit was estimated as 3.3 μg mL(-1). Meanwhile, selectivity experiments demonstrated that imprinted gel had a high affinity to carnosine in the presence of close structural analogues (interferrants). PMID:25491843

  5. Molecular mechanism of MLL PHD3 and RNA recognition by the Cyp33 RRM domain

    PubMed Central

    Hom, Robert A.; Chang, Pei-Yun; Roy, Siddhartha; Musselman, Catherine A.; Glass, Karen C.; Selezneva, Anna I.; Gozani, Or; Ismagilov, Rustem F.; Cleary, Michael L.; Kutateladze, Tatiana G.

    2010-01-01

    Summary The nuclear protein Cyclophilin 33 (Cyp33) is a peptidyl-prolyl cis-trans isomerase that catalyzes cis-trans isomerization of the peptide bond preceding a proline and promotes folding and conformational changes in folded and unfolded proteins. The N-terminal RRM domain of Cyp33 has been found to associate with the third plant homeodomain (PHD3) finger of the Mixed Lineage Leukemia (MLL) proto-oncoprotein and a poly-A RNA sequence. Here, we report a 1.9 Å resolution crystal structure of the RRM domain of Cyp33 and describe the molecular mechanism of PHD3 and RNA recognition. The Cyp33 RRM domain folds into a five-stranded antiparallel β-sheet and two α-helices. The RRM domain but not the catalytic module of Cyp33 binds strongly to PHD3, exhibiting a 2 μM affinity as measured by Isothermal Titration Calorimetry (ITC). NMR chemical shift perturbation (CSP) analysis and dynamics data reveal that the β strands and the β2–β3 loop of the RRM domain are involved in the interaction with PHD3. Mutations in the PHD3-binding site or deletions in the β 2/β 3 loop lead to a significantly reduced affinity or abrogation of the interaction. The RNA-binding pocket of the Cyp33 RRM domain, mapped based on NMR CSP and mutagenesis, partially overlaps with the PHD3-binding site, and RNA association is abolished in the presence of MLL PHD3. Full-length Cyp33 acts as a negative regulator of MLL-induced transcription and reduces the expression levels of MLL target genes MEIS1 and HOXA9. Together, these in vitro and in vivo data provide insight into the multiple functions of Cyp33 RRM and suggest a Cyp33-dependent mechanism for regulating the transcriptional activity of MLL. PMID:20460131

  6. Probability model for molecular recognition in biological receptor repertoires: significance to the olfactory system.

    PubMed Central

    Lancet, D; Sadovsky, E; Seidemann, E

    1993-01-01

    A generalized phenomenological model is presented for stereospecific recognition between biological receptors and their ligands. We ask what is the distribution of binding constants psi(K) between an arbitrary ligand and members of a large receptor repertoire, such as immunoglobulins or olfactory receptors. For binding surfaces with B potential subsite and S different types of subsite configurations, the number of successful elementary interactions obeys a binomial distribution. The discrete probability function psi(K) is then derived with assumptions on alpha, the free energy contribution per elementary interaction. The functional form of psi(K) may be universal, although the parameter values could vary for different ligand types. An estimate of the parameter values of psi(K) for iodovanillin, an analog of odorants and immunological haptens, is obtained by equilibrium dialysis experiments with nonimmune antibodies. Based on a simple relationship, predicted by the model, between the size of a receptor repertoire and its average maximal affinity toward an arbitrary ligand, the size of the olfactory receptor repertoire (Nolf) is calculated as 300-1000, in very good agreement with recent molecular biological studies. A very similar estimate, Nolf = 500, is independently derived by relating a theoretical distribution of maxima for psi(K) with published human olfactory threshold variations. The present model also has implications to the question of olfactory coding and to the analysis of specific anosmias, genetic deficits in perceiving particular odorants. More generally, the proposed model provides a better understanding of ligand specificity in biological receptors and could help in understanding their evolution. PMID:8475121

  7. Recognition Properties and Competitive Assays of a Dual Dopamine/Serotonin Selective Molecularly Imprinted Polymer

    PubMed Central

    Suedee, Roongnapa; Seechamnanturakit, Vatcharee; Suksuwan, Acharee; Canyuk, Bhutorn

    2008-01-01

    A molecularly imprinted polymer (MIP) with dual dopamine/serotonin-like binding sites (DS-MIP) was synthesized for use as a receptor model of study the drug-interaction of biological mixed receptors at a molecular level. The polymer material was produced using methacrylic acid (MAA) and acrylamide (ACM) as functional monomers, N,N′-methylene bisacrylamide (MBAA) as cross-linker, methanol/water mixture (4:1, v/v) as porogen and a mixture of dopamine (D) and serotonin (S) as templates. The prepared DS-MIP exhibited the greatest rebinding of the template(s) in aqueous methanol solution with decreased recognition in acetonitrile, water and methanol solvent. The binding affinity and binding capacity of DS-MIP with S were found to be higher than those of DS-MIP with D. The selectivity profiles of DS-MIP suggest that the D binding site of DS-MIP has sufficient integrity to discriminate between species of non-optimal functional group orientation, whilst the S binding site of DS-MIP is less selective toward species having structural features and functional group orientations different from S. The ligand binding activities of a series of ergot derivatives (ergocryptine, ergocornine, ergocristine, ergonovine, agroclavine, pergolide and terguride) have been studied with the DS-MIP using a competitive ligand binding assay protocol. The binding affinities of DS-MIP were demonstrated in the micro- or submicro-molar range for a series of ergot derivatives, whereas the binding affinities were considerably greater to natural receptors derived from the rat hypothalamus. The DS-MIP afforded the same pattern of differentiation as the natural receptors, i.e. affinity for the clavines > lysergic acid derivatives > ergopeptines. The results suggest that the discrimination for the ergot derivatives by the dopamine and serotonin sites of DS-MIP is due to the structural features and functional orientation of the phenylethylamine and indolylethylamine entities at the binding sites, and the

  8. Development and molecular characterization of polymeric micro-nanofibrous scaffold of a defined 3-D niche for in vitro chemosensitivity analysis against acute myeloid leukemia cells

    PubMed Central

    Nair, Maya S; Mony, Ullas; Menon, Deepthy; Koyakutty, Manzoor; Sidharthan, Neeraj; Pavithran, Keechilat; Nair, Shantikumar V; Menon, Krishnakumar N

    2015-01-01

    Standard in vitro drug testing employs 2-D tissue culture plate systems to test anti-leukemic drugs against cell adhesion-mediated drug-resistant leukemic cells that harbor in 3-D bone marrow microenvironments. This drawback necessitates the fabrication of 3-D scaffolds that have cell adhesion-mediated drug-resistant properties similar to in vivo niches. We therefore aimed at exploiting the known property of polyurethane (PU)/poly-l-lactic acid (PLLA) in forming a micro-nanofibrous structure to fabricate unique, not presented before, as far as we are aware, 3-D micro-nanofibrous scaffold composites using a thermally induced phase separation technique. Among the different combinations of PU/PLLA composites generated, the unique PU/PLLA 60:40 composite displayed micro-nanofibrous morphology similar to decellularized bone marrow with increased protein and fibronectin adsorption. Culturing of acute myeloid leukemia (AML) KG1a cells in FN-coated PU/PLLA 60:40 shows increased cell adhesion and cell adhesion-mediated drug resistance to the drugs cytarabine and daunorubicin without changing the original CD34+/CD38−/CD33− phenotype for 168 hours compared to fibronectin tissue culture plate systems. Molecularly, as seen in vivo, increased chemoresistance is associated with the upregulation of anti-apoptotic Bcl2 and the cell cycle regulatory protein p27Kip1 leading to cell growth arrest. Abrogation of Bcl2 activity by the Bcl2-specific inhibitor ABT 737 led to cell death in the presence of both cytarabine and daunorubicin, demonstrating that the cell adhesion-mediated drug resistance induced by Bcl2 and p27Kip1 in the scaffold was similar to that seen in vivo. These results thus show the utility of a platform technology, wherein drug testing can be performed before administering to patients without the necessity for stromal cells. PMID:26028971

  9. Core-shell molecularly imprinted polymer nanoparticles with assistant recognition polymer chains for effective recognition and enrichment of natural low-abundance protein.

    PubMed

    Liu, Dejing; Yang, Qian; Jin, Susu; Song, Yingying; Gao, Junfei; Wang, Ying; Mi, Huaifeng

    2014-02-01

    Core-shell molecular imprinting of nanomaterials overcomes difficulties with template transfer and achieves higher binding capacities for macromolecular imprinting, which are more important to the imprinting of natural low-abundance proteins from cell extracts. In the present study, a novel strategy of preparing core-shell nanostructured molecularly imprinted polymers (MIPs) was developed that combined the core-shell approach with assistant recognition polymer chains (ARPCs). Vinyl-modified silica nanoparticles were used as support and ARPCs were used as additional functional monomers. Immunoglobulin heavy chain binding protein (BiP) from the endoplasmic reticulum (ER) was chosen as the model protein. The cloned template protein BiP was selectively assembled with ARPCs from their library, which contained numerous limited-length polymer chains with randomly distributed recognition and immobilization sites. The resulting complex was copolymerized onto the surface of vinyl-modified silica nanoparticles under low concentrations of the monomers. After template removal, core-shell-structured nanoparticles with a thin imprinted polymer layer were produced. The particles demonstrated considerably high adsorption capacity, fast adsorption kinetics and selective binding affinities toward the template BiP. Furthermore, the synthesized MIP nanoparticles successfully isolated cloned protein BiP from protein mixtures and highly enriched BiP from an ER extract containing thousands of kinds of proteins. The enrichment reached 115-fold and the binding capacity was 5.4 μg g(-1), which were higher than those achieved by using traditional MIP microspheres. The advantageous properties of MIP nanoparticles hold promise for further practical applications in biology, such as protein analysis and purification. PMID:24140608

  10. Molecular basis of the structural stability of a Top7-based scaffold at extreme pH and temperature conditions.

    PubMed

    Soares, Thereza A; Boschek, Curt B; Apiyo, David; Baird, Cheryl; Straatsma, T P

    2010-06-01

    The development of stable biomolecular scaffolds that can tolerate environmental extremes has considerable potential for industrial and defense-related applications. However, most natural proteins are not sufficiently stable to withstand non-physiological conditions. We have recently engineered the de novo designed Top7 protein to specifically recognize the glycoprotein CD4 by insertion of an eight-residue loop. The engineered variant exhibited remarkable stability under chemical and thermal denaturation conditions. In the present study, far-UV CD spectroscopy and explicit-solvent MD simulations are used to investigate the structural stability of Top7 and the engineered variant under extreme conditions of temperature and pH. Circular dichroism measurements suggest that the engineered variant Top7(CB1), like Top7, retains its structure at high temperatures. Changes in CD spectra suggest that there are minor structural rearrangements between neutral and acidic environments for both proteins but that these do not make the proteins less stable at high temperatures. The anti-parallel beta-sheet is well conserved within the timescale simulated whereas there is a decrease of helical content when low pH and high-temperature conditions are combined. Concerted alanine mutations along the alpha-helices of the engineered Top7 variant did not revert this trend when at pH 2 and 400K. The structural resilience of the anti-parallel beta-sheet suggests that the protein scaffold can accommodate varying sequences. The robustness of the Top7 scaffold under extreme conditions of pH and temperature and its amenability to production in inexpensive bacterial expression systems reveal great potential for novel biotechnological applications. PMID:20185346

  11. Water-compatible temperature and magnetic dual-responsive molecularly imprinted polymers for recognition and extraction of bisphenol A.

    PubMed

    Wu, Xiaqing; Wang, Xiaoyan; Lu, Wenhui; Wang, Xinran; Li, Jinhua; You, Huiyan; Xiong, Hua; Chen, Lingxin

    2016-02-26

    Versatile molecularly imprinted polymers (MIPs) have been widely applied to various sample matrices, however, molecular recognition in aqueous media is still difficult. Stimuli-responsive MIPs have received increasing attentions due to their unique feature that the molecular recognition is regulated by specific external stimuli. Herein, water-compatible temperature and magnetic dual-responsive MIPs (WC-TMMIPs) with hydrophilic brushes were prepared via reversible addition-fragmentation chain transfer precipitation polymerization for reversible and selective recognition and extraction of bisphenol A (BPA). Transmission electron microscopy (TEM), Fourier transform infrared spectrometer (FT-IR) and vibrating sample magnetometry (VSM) as characterization methods were used to examine the successful synthesis of polymers, and the resultant WC-TMMIPs showed excellent thermosensitivity and simple rapid magnetic separation. Controlled adsorption and release of BPA by temperature regulation were investigated systematically, and the maximum adsorption and removal efficiency toward BPA in aqueous solutions were attained at 35 °C and 45 °C, respectively, as well as a good recoverability was exhibited with the precision less than 5% through five adsorption-desorption cycles. Phenolic structural analogs were tested and good recognition specificity for BPA was displayed. Accordingly, the WC-TMMIPs were employed as adsorbents for magnetic solid-phase extraction (MSPE) and packed SPE of BPA from seawater samples. Using the two modes followed by HPLC-UV determination, excellent linearity was attained in the range of 0.1-14.5 μM and 1.3-125 nM, with low detection limits of 0.02 μM and 0.18 nM, respectively. Satisfactory recoveries for spiked seawater samples were achieved ranging from 86.3-103.5% and 96.2-104.3% with RSD within 2.12-4.33%. The intelligent WC-TMMIPs combining water-compatibility, molecular recognition, magnetic separation, and temperature regulation proved

  12. Synthesis, antimicrobial activity and molecular docking of novel tetracyclic scaffolds incorporating a flavonoid framework with medium sized oxygen heterocycles.

    PubMed

    Dongamanti, Ashok; Aamate, Vikas Kumar; Devulapally, Mohan Gandhi; Gundu, Srinivas; Kotni, Meena Kumari; Manga, Vijjulatha; Balasubramanian, Sridhar; Ernala, Prasad

    2015-02-15

    A convenient approach for the synthesis of novel tetracyclic scaffolds incorporating a flavonoid framework with medium sized heterocyclic rings (eight-, nine-, ten- and eleven-membered rings) containing two oxygen atoms from flavonols through alkylation using different dibromoalkanes was described. The synthesized compounds were established based on the spectral data and X-ray crystal structure for 6c. The synthesized compounds were evaluated for their in vitro antimicrobial activity. Docking studies were carried out for most active two compounds 6f and 6i. PMID:25592711

  13. Molecular construction of HIV-gp120 discontinuous epitope mimics by assembly of cyclic peptides on an orthogonal alkyne functionalized TAC-scaffold.

    PubMed

    Werkhoven, P R; Elwakiel, M; Meuleman, T J; Quarles van Ufford, H C; Kruijtzer, J A W; Liskamp, R M J

    2016-01-14

    Mimics of discontinuous epitopes of for example bacterial or viral proteins may have considerable potential for the development of synthetic vaccines, especially if conserved epitopes can be mimicked. However, due to the structural complexity and size of discontinuous epitopes molecular construction of these mimics remains challeging. We present here a convergent route for the assembly of discontinuous epitope mimics by successive azide alkyne cycloaddition on an orthogonal alkyne functionalized scaffold. Here the synthesis of mimics of the HIV gp120 discontinuous epitope that interacts with the CD4 receptor is described. The resulting protein mimics are capable of inhibition of the gp120-CD4 interaction. The route is convergent, robust and should be applicable to other discontinuous epitopes. PMID:26552661

  14. Optimized solid phase-assisted synthesis of dendrons applicable as scaffolds for radiolabeled bioactive multivalent compounds intended for molecular imaging.

    PubMed

    Fischer, Gabriel; Wängler, Björn; Wängler, Carmen

    2014-01-01

    Dendritic structures, being highly homogeneous and symmetric, represent ideal scaffolds for the multimerization of bioactive molecules and thus enable the synthesis of compounds of high valency which are e.g., applicable in radiolabeled form as multivalent radiotracers for in vivo imaging. As the commonly applied solution phase synthesis of dendritic scaffolds is cumbersome and time-consuming, a synthesis strategy was developed that allows for the efficient assembly of acid amide bond-based highly modular dendrons on solid support via standard Fmoc solid phase peptide synthesis protocols. The obtained dendritic structures comprised up to 16 maleimide functionalities and were derivatized on solid support with the chelating agent DOTA. The functionalized dendrons furthermore could be efficiently reacted with structurally variable model thiol-bearing bioactive molecules via click chemistry and finally radiolabeled with 68Ga. Thus, this solid phase-assisted dendron synthesis approach enables the fast and straightforward assembly of bioactive multivalent constructs for example applicable as radiotracers for in vivo imaging with Positron Emission Tomography (PET). PMID:24871573

  15. Toward a Molecular Lego Approach for the Diversity-Oriented Synthesis of Cyclodextrin Analogues Designed as Scaffolds for Multivalent Systems.

    PubMed

    Lepage, Mathieu L; Schneider, Jérémy P; Bodlenner, Anne; Compain, Philippe

    2015-11-01

    A modular strategy has been developed to access a diversity of cyclic and acyclic oligosaccharide analogues designed as prefunctionalized scaffolds for the synthesis of multivalent ligands. This convergent approach is based on bifunctional sugar building blocks with two temporarily masked functionalities that can be orthogonally activated to perform Cu(I)-catalyzed azide-alkyne cycloaddition reactions (CuAAC). The reducing end is activated as a glycosyl azide and masked as a 1,6-anhydro sugar, while the nonreducing end is activated as a free alkyne and masked as a triethylsilyl-alkyne. Following a cyclooligomerization approach, the first examples of close analogues of cyclodextrins composed of d-glucose residues and triazole units bound together through α-(1,4) linkages were obtained. The cycloglucopyranoside analogue containing four sugar units was used as a template to prepare multivalent systems displaying a protected d-mannose derivative or an iminosugar by way of CuAAC. On the other hand, the modular approach led to acyclic alkyne-functionalized scaffolds of a controlled size that were used to synthesize multivalent iminosugars. PMID:26439895

  16. “One Ring to Bind Them All”—Part I: The Efficiency of the Macrocyclic Scaffold for G-Quadruplex DNA Recognition

    PubMed Central

    Monchaud, David; Granzhan, Anton; Saettel, Nicolas; Guédin, Aurore; Mergny, Jean-Louis; Teulade-Fichou, Marie-Paule

    2010-01-01

    Macrocyclic scaffolds are particularly attractive for designing selective G-quadruplex ligands essentially because, on one hand, they show a poor affinity for the “standard” B-DNA conformation and, on the other hand, they fit nicely with the external G-quartets of quadruplexes. Stimulated by the pioneering studies on the cationic porphyrin TMPyP4 and the natural product telomestatin, follow-up studies have developed, rapidly leading to a large diversity of macrocyclic structures with remarkable-quadruplex binding properties and biological activities. In this review we summarize the current state of the art in detailing the three main categories of quadruplex-binding macrocycles described so far (telomestatin-like polyheteroarenes, porphyrins and derivatives, polyammonium cyclophanes), and in addressing both synthetic issues and biological aspects. PMID:20725629

  17. Scaffolded biology.

    PubMed

    Minelli, Alessandro

    2016-09-01

    Descriptions and interpretations of the natural world are dominated by dichotomies such as organism vs. environment, nature vs. nurture, genetic vs. epigenetic, but in the last couple of decades strong dissatisfaction with those partitions has been repeatedly voiced and a number of alternative perspectives have been suggested, from perspectives such as Dawkins' extended phenotype, Turner's extended organism, Oyama's Developmental Systems Theory and Odling-Smee's niche construction theory. Last in time is the description of biological phenomena in terms of hybrids between an organism (scaffolded system) and a living or non-living scaffold, forming unit systems to study processes such as reproduction and development. As scaffold, eventually, we can define any resource used by the biological system, especially in development and reproduction, without incorporating it as happens in the case of resources fueling metabolism. Addressing biological systems as functionally scaffolded systems may help pointing to functional relationships that can impart temporal marking to the developmental process and thus explain its irreversibility; revisiting the boundary between development and metabolism and also regeneration phenomena, by suggesting a conceptual framework within which to investigate phenomena of regular hypermorphic regeneration such as characteristic of deer antlers; fixing a periodization of development in terms of the times at which a scaffolding relationship begins or is terminated; and promoting plant galls to legitimate study objects of developmental biology. PMID:27287514

  18. Supramolecular polymers constructed from macrocycle-based host-guest molecular recognition motifs.

    PubMed

    Dong, Shengyi; Zheng, Bo; Wang, Feng; Huang, Feihe

    2014-07-15

    /physical properties, including stimuli responsiveness, self-healing, and environmental adaptation. It has been reported that macrocycle-based supramolecular polymers can respond to pH change, photoirradition, anions, cations, temperature, and solvent. Macrocycle-based supramolecular polymers have been prepared in solution, in gel, and in the solid state. Furthermore, the solvent has a very important influence on the formation of these supramolecular polymers. Crown ether- and pillararene-based supramolecular polymers have mainly formed in organic solvents, such as chloroform, acetone, and acetonitrile, while cyclodextrin- and cucurbituril-based supramolecular polymerizations have been usually observed in aqueous solutions. For calixarenes, both organic solvents and water have been used as suitable media for supramolecular polymerization. With the development of supramolecular chemistry and polymer science, various methods, such as nuclear magnetic resonance spectroscopy, X-ray techniques, electron microscopies, and theoretical calculation and computer simulation, have been applied for characterizing supramolecular polymers. The fabrication of macrocycle-based supramolecular polymers has become a currently hot research topic. In this Account, we summarize recent results in the investigation of supramolecular polymers constructed from macrocycle-based host-guest molecular recognition motifs. These supramolecular polymers are classified based on the different macrocycles used in them. Their monomer design, structure control, stimuli-responsiveness, and applications in various areas are discussed, and future research directions are proposed. It is expected that the development of supramolecular polymers will not only change the way we live and work but also exert significant influence on scientific research. PMID:24684594

  19. Influence of mobile phase composition and cross-linking density on the enantiomeric recognition properties of molecularly imprinted polymers.

    PubMed

    Yu, C; Mosbach, K

    2000-08-01

    A series of experiments were conducted to investigate elements which affect the enantiomeric recognition properties of molecularly imprinted polymers (MIPs) in the HPLC mode. Our results show that the recognition properties of MIPs are greatly influenced by the mobile phase used. For a polymer prepared in acetonitrile, a good enantiomeric separation was observed when acetonitrile-based mobile phase was used, when the mobile phase was changed to chloroform-based, no enantiomeric recognition was observed although the sample molecule was retarded. This indicates that the specific co-operative binding interactions between the functional groups at the imprinted polymer's recognition sites and the sample molecule were considerably disrupted and only non-specific interactions remained. When the mobile phase was changed back to acetonitrile-based, the recognition was regained. In contrast, for polymers prepared in chloroform, chloroform-based mobile phase gave much better separation than acetonitrile-based mobile phase. When other solvents were tested, significant solvent effects were generally observed. Based on these observations, the recognition properties of the methacrylic acid (MAA)-co-ethylene glycol dimethacrylate (EGDMA) polymers were reinvestigated, and the results show that by simply using an optimised mobile phase system, significantly improved recognition over previously reported results was observed. For a polymer made against Cbz-L-Trp, 100 microg of Cbz-D,L-Trp was separated with a separation factor (alpha) of 4.23 and a resolution (Rs) of 3.87, whereas in the previous report, 10 microg of Cbz-D,L-Trp was only separated with alpha = 1.67 and Rs = 0.1. It is generally realised that the imprinted polymer's recognition property is also very much influenced by the nature of the polymer network. It was shown that the recognition decreased with a decrease in the apparent degree of cross-linking (molar percentage of cross-linker in the polymerisation mixture

  20. Immobilization of oligonucleotides onto zirconia-modified filter paper and specific molecular recognition.

    PubMed

    Xiao, Wei; Huang, Jianguo

    2011-10-18

    A morphologically complex cellulosic substance (e.g., commercial filter paper) was employed as a substrate for DNA immobilization and successive recognition. A uniform ultrathin zirconia gel film was first deposited on each cellulose nanofiber in bulk filter paper by a facile sol-gel process. Relying on the large surface area of filter paper and the strong affinity of zirconia for the phosphate group, terminal-phosphate probe DNA was abundantly immobilized on the zirconia-modified filter paper so as to convert the composite to a biofunctional material for the sensitive and repetitive recognition of the corresponding complementary target DNA on the nanomolar level. By contrast, in spite of the viability of the immobilization of the probe DNA and the recognition of target DNA on the quartz plate, the amount of captured probe DNA or recognized target DNA on such a flat substrate was much less than that captured or recognized on filter paper, resulting in a relatively insensitive recognition event. Moreover, control experiments on bare filter paper (without a zirconia nanocoating) suggested that the zirconia gel film was essential to probe DNA immobilization and subsequent target DNA recognition. PMID:21905718

  1. Molecular Recognition of PTS-1 Cargo Proteins by Pex5p: Implications for Protein Mistargeting in Primary Hyperoxaluria

    PubMed Central

    Mesa-Torres, Noel; Tomic, Nenad; Albert, Armando; Salido, Eduardo; Pey, Angel L.

    2015-01-01

    Peroxisomal biogenesis and function critically depends on the import of cytosolic proteins carrying a PTS1 sequence into this organelle upon interaction with the peroxin Pex5p. Recent structural studies have provided important insights into the molecular recognition of cargo proteins by Pex5p. Peroxisomal import is a key feature in the pathogenesis of primary hyperoxaluria type 1 (PH1), where alanine:glyoxylate aminotransferase (AGT) undergoes mitochondrial mistargeting in about a third of patients. Here, we study the molecular recognition of PTS1 cargo proteins by Pex5p using oligopeptides and AGT variants bearing different natural PTS1 sequences, and employing an array of biophysical, computational and cell biology techniques. Changes in affinity for Pex5p (spanning over 3–4 orders of magnitude) reflect different thermodynamic signatures, but overall bury similar amounts of molecular surface. Structure/energetic analyses provide information on the contribution of ancillary regions and the conformational changes induced in Pex5p and the PTS1 cargo upon complex formation. Pex5p stability in vitro is enhanced upon cargo binding according to their binding affinities. Moreover, we provide evidence that the rational modulation of the AGT: Pex5p binding affinity might be useful tools to investigate mistargeting and misfolding in PH1 by pulling the folding equilibria towards the native and peroxisomal import competent state. PMID:25689234

  2. Spectroscopy, NMR and DFT studies on molecular recognition of crown ether bridged chiral heterotrinuclear salen Zn(II) complex

    NASA Astrophysics Data System (ADS)

    Gao, Feng; Ruan, Wen-Juan; Chen, Jia-Mei; Zhang, Ying-Hui; Zhu, Zhi-Ang

    2005-12-01

    A barium-containing crown ether bridged chiral heterotrinuclear salen Zn(II) complex BaZn 2L(ClO 4) 2, where L is a folded dinuclear chiral ( R, R)-salen ligand, has been synthesized and characterized by elemental analysis, 1H NMR, UV-vis, IR, circular dichroism (CD) spectra, and mass spectra. As a folded dinuclear chiral host, its recognition with achiral guests (imidazole derivatives), rigid bidentate guest (1,4-diazobicyclo[2,2,2]octane, DABCO) and chiral guests (amino acid methyl esters) was investigated by means of UV-vis spectrophotometric titration, CD spectra. The association constants of D-amino acid methyl esters are found to be higher than those of their L-enantiomer. The sandwich-type binding of BaZn 2L(ClO 4) 2-DABCO supramolecular assembly was specially studied via 1H NMR titration and 1H ROESY. To understand the recognition on molecular level, density functional theory (DFT) calculations on B3LYP/LanL2DZ were performed on the minimal energy conformations of host, guests, and host-guest complexes. The minimal energy conformations were obtained by molecular mechanics (MM) optimization and molecular dynamics (MD) simulation. The results of single point energy, HOMO energy, and charges transfer were analyzed. The results of theoretical calculations are in good agreement with the experimental data.

  3. Supramolecular Alternating Polymer from Crown Ether and Pillar[5]arene-Based Double Molecular Recognition for Preparation of Hierarchical Materials.

    PubMed

    Li, Hui; Fan, Xiaodong; Qi, Miao; Yang, Zhen; Zhang, Haitao; Tian, Wei

    2016-01-01

    A novel supramolecular alternating polymer is constructed based on double molecular recognition events of benzo-21-crown-7 with a secondary ammonium salt and of pillar[5]arene with a neutral guest. The resulting polymer is utilized to prepare hierarchical materials with different dimensionalities for the first time. These materials included zero-dimensional spherical aggregates, one-dimensional nanofibers, two-dimensional microstructured films, and three-dimensional ordered glue. This development will be helpful for designing and preparing supramolecular hierarchical materials with different dimensionalities. PMID:26555439

  4. Dynamics and recognition within a protein–DNA complex: a molecular dynamics study of the SKN-1/DNA interaction

    PubMed Central

    Etheve, Loïc; Martin, Juliette; Lavery, Richard

    2016-01-01

    Molecular dynamics simulations of the Caenorhabditis elegans transcription factor SKN-1 bound to its cognate DNA site show that the protein–DNA interface undergoes significant dynamics on the microsecond timescale. A detailed analysis of the simulation shows that movements of two key arginine side chains between the major groove and the backbone of DNA generate distinct conformational sub-states that each recognize only part of the consensus binding sequence of SKN-1, while the experimentally observed binding specificity results from a time-averaged view of the dynamic recognition occurring within this complex. PMID:26721385

  5. Selectivity of pyoverdine recognition by the FpvA receptor of Pseudomonas aeruginosa from molecular dynamics simulations.

    PubMed

    Bouvier, Benjamin; Cézard, Christine; Sonnet, Pascal

    2015-07-21

    The Gram-negative bacterium Pseudomonas aeruginosa, a ubiquitous human opportunistic pathogen, has developed resistances to multiple antibiotics. It uses its primary native siderophore, pyoverdine, to scavenge the iron essential to its growth in the outside medium and transport it back into its cytoplasm. The FpvA receptor on the bacterial outer membrane recognizes and internalizes pyoverdine bearing its iron payload, but can also bind pyoverdines from other Pseudomonads or synthetic analogues. Pyoverdine derivatives could therefore be used as vectors to deliver antibiotics into the bacterium. In this study, we use molecular dynamics and free energy calculations to characterize the mechanisms and thermodynamics of the recognition of the native pyoverdines of P. aeruginosa and P. fluorescens by FpvA. Based on these results, we delineate the features that pyoverdines with high affinity for FpvA should possess. In particular, we show that (i) the dynamics and interaction of the unbound pyoverdines with water should be optimized with equal care as the interface contacts in the complex with FpvA; (ii) the C-terminal extremity of the pyoverdine chain, which appears to play no role in the bound complex, is involved in the intermediate stages of recognition; and (iii) the length and cyclicity of the pyoverdine chain can be used to fine-tune the kinetics of the recognition mechanism. PMID:26098682

  6. Atomic-Scale Insight into Tautomeric Recognition, Separation, and Interconversion of Guanine Molecular Networks on Au(111).

    PubMed

    Zhang, Chi; Xie, Lei; Wang, Likun; Kong, Huihui; Tan, Qinggang; Xu, Wei

    2015-09-16

    Although tautomerization may directly affect the chemical or biological properties of molecules, real-space investigation on the tautomeric behaviors of organic molecules in a larger area of molecular networks has been scarcely reported. In this paper, we choose guanine (G) molecule as a model system. From the interplay of high-resolution scanning tunneling microscopy (STM) imaging and density functional theory (DFT) calculations, we have successfully achieved the tautomeric recognition, separation, and interconversion of G molecular networks (formed by two tautomeric forms G/9H and G/7H) with the aid of NaCl on the Au(111) surface in ultrahigh vacuum (UHV) conditions. Our results may serve as a prototypical system to provide important insights into tautomerization related issues, which should be intriguing to biochemistry, pharmaceutics, and other related fields. PMID:26322860

  7. Single cell molecular recognition of migrating and invading tumor cells using a targeted fluorescent probe to receptor PTPmu.

    PubMed

    Burden-Gulley, Susan M; Qutaish, Mohammed Q; Sullivant, Kristin E; Tan, Mingqian; Craig, Sonya E L; Basilion, James P; Lu, Zheng-Rong; Wilson, David L; Brady-Kalnay, Susann M

    2013-04-01

    Detection of an extracellular cleaved fragment of a cell-cell adhesion molecule represents a new paradigm in molecular recognition and imaging of tumors. We previously demonstrated that probes that recognize the cleaved extracellular domain of receptor protein tyrosine phosphatase mu (PTPmu) label human glioblastoma brain tumor sections and the main tumor mass of intracranial xenograft gliomas. In this article, we examine whether one of these probes, SBK2, can label dispersed glioma cells that are no longer connected to the main tumor mass. Live mice with highly dispersive glioma tumors were injected intravenously with the fluorescent PTPmu probe to test the ability of the probe to label the dispersive glioma cells in vivo. Analysis was performed using a unique three-dimensional (3D) cryo-imaging technique to reveal highly migratory and invasive glioma cell dispersal within the brain and the extent of colabeling by the PTPmu probe. The PTPmu probe labeled the main tumor site and dispersed cells up to 3.5 mm away. The cryo-images of tumors labeled with the PTPmu probe provide a novel, high-resolution view of molecular tumor recognition, with excellent 3D detail regarding the pathways of tumor cell migration. Our data demonstrate that the PTPmu probe recognizes distant tumor cells even in parts of the brain where the blood-brain barrier is likely intact. The PTPmu probe has potential translational significance for recognizing tumor cells to facilitate molecular imaging, a more complete tumor resection and to serve as a molecular targeting agent to deliver chemotherapeutics to the main tumor mass and distant dispersive tumor cells. PMID:22987116

  8. Multivalent display of proteins on viral nanoparticles using molecular recognition and chemical ligation strategies.

    PubMed

    Venter, P Arno; Dirksen, Anouk; Thomas, Diane; Manchester, Marianne; Dawson, Philip E; Schneemann, Anette

    2011-06-13

    Multivalent display of heterologous proteins on viral nanoparticles forms a basis for numerous applications in nanotechnology, including vaccine development, targeted therapeutic delivery, and tissue-specific bioimaging. In many instances, precise placement of proteins is required for optimal functioning of the supramolecular assemblies, but orientation- and site-specific coupling of proteins to viral scaffolds remains a significant technical challenge. We have developed two strategies that allow for controlled attachment of a variety of proteins on viral particles using covalent and noncovalent principles. In one strategy, an interaction between domain 4 of anthrax protective antigen and its receptor was used to display multiple copies of a target protein on virus-like particles. In the other, expressed protein ligation and aniline-catalyzed oximation was used to display covalently a model protein. The latter strategy, in particular, yielded nanoparticles that induced potent immune responses to the coupled protein, suggesting potential applications in vaccine development. PMID:21545187

  9. Silylated melamine and cyanuric acid as precursors for imprinted and hybrid silica materials with molecular recognition properties.

    PubMed

    Arrachart, Guilhem; Carcel, Carole; Trens, Philippe; Moreau, Jöel J E; Wong Chi Man, Michel

    2009-06-15

    Two monotrialkoxysilylated compounds that consist of complementary fragments of melamine (M) and cyanuric acid (CA) have been synthesised. The molecular recognition properties of the M and CA fragments through complementary hydrogen bonds (DAD and ADA; D=donor, A=acceptor) are the key factor used to direct the formation of hybrid silica materials by using a sol-gel process. These materials were synthesised following two methods: First, an organo-bridged silsesquioxane was obtained by the hydrolysis of the two complementary monotrialkoxysilylated melamine and cyanuric acid derivatives, with fluoride ions as a catalyst. The hydrogen-bonding interactions between the two organic fragments are responsible for the formation of the bridging unit. The transcription of the assembly into the hybrid material was characterised and evidenced by solid-state NMR (29Si, 13C) and FTIR spectroscopic experiments. Second, the molecular recognition was exploited to synthesise an imprinted hybrid silica. This material was prepared by co-condensation of tetraethyl orthosilicate (TEOS) with the monosilylated cyanuric acid derivative (CA) templated by nonsilylated melamine. The melamine template was completely removed by treating the solid material with hydrochloric acid. The reintroduction of the template was performed by treating the resulting material with an aqueous suspension of melamine. These steps were monitored and analysed by several techniques, such as solid-state NMR (29Si, 13C) and FTIR spectroscopic analysis and nitrogen adsorption-desorption isotherms. PMID:19440996

  10. Applying pattern recognition methods to analyze the molecular properties of a homologous series of nitrogen mustard agents.

    PubMed

    Bartzatt, Ronald; Donigan, Laura

    2006-01-01

    The purpose of this research was to analyze the pharmacological properties of a homologous series of nitrogen mustard (N-mustard) agents formed after inserting 1 to 9 methylene groups (-CH2-) between 2 -N(CH2CH2Cl)2 groups. These compounds were shown to have significant correlations and associations in their properties after analysis by pattern recognition methods including hierarchical classification, cluster analysis, nonmetric multi-dimensional scaling (MDS), detrended correspondence analysis, K-means cluster analysis, discriminant analysis, and self-organizing tree algorithm (SOTA) analysis. Detrended correspondence analysis showed a linear-like association of the 9 homologs, and hierarchical classification showed that each homolog had great similarity to at least one other member of the series-as did cluster analysis using paired-group distance measure. Nonmetric multi-dimensional scaling was able to discriminate homologs 2 and 3 (by number of methylene groups) from homologs 4, 5, and 6 as a group, and from homologs 7, 8, and 9 as a group. Discriminant analysis, K-means cluster analysis, and hierarchical classification distinguished the high molecular weight homologs from low molecular weight homologs. As the number of methylene groups increased the aqueous solubility decreased, dermal permeation coefficient increased, Log P increased, molar volume increased, parachor increased, and index of refraction decreased. Application of pattern recognition methods discerned useful interrelationships within the homologous series that will determine specific and beneficial clinical applications for each homolog and methods of administration. PMID:16796353

  11. Broad-Specificity Chemiluminescence Enzyme Immunoassay for (Fluoro)quinolones: Hapten Design and Molecular Modeling Study of Antibody Recognition.

    PubMed

    Zeng, Haopeng; Chen, Jiahong; Zhang, Chijian; Huang, Xin-An; Sun, Yuanming; Xu, Zhenlin; Lei, Hongtao

    2016-04-01

    On the basis of the structural features of (fluoro)quinolones (FQs), pazufloxacin was first used as a generic immunizing hapten to raise a broad-specificity antibody. The obtained polyclonal antibody exhibited broad cross-reactivity ranging from 5.19% to 478.77% with 21 FQs. Furthermore, the antibody was able to recognize these FQs below their maximum residue limits (MRLs) in an indirect competitive chemiluminescence enzyme immunoassay (ic-CLEIA), with the limit of detection (LOD) ranging from 0.10 to 33.83 ng/mL. For simply pretreated milk samples with spiked FQs, the ic-CLEIA exhibited an excellent recovery with a range of 84.6-106.9% and an acceptable coefficient of variation below 15%, suggesting its suitability and reliability for the use of a promising tool to detect FQs. Meanwhile, comparative molecular field analysis (CoMFA) and comparative molecular similarity indices analysis (CoMSIA) models, with statistically significant correlation coefficients (q(2)CoMFA = 0.559, r(2)CoMFA = 0.999; q(2)CoMSIA = 0.559, r(2)CoMSIA = 0.994), were established to investigate the antibody recognition mechanism. These two models revealed that in the antibody, the active cavity binding FQs' 7-position substituents worked together with another cavity (binding FQs' 1-position groups) to crucially endow the high cross-reactivity. This investigation will be significant for better exploring the recognition mechanism and for designing new haptens. PMID:26976361

  12. Molecular Basis for LLT1 Protein Recognition by Human CD161 Protein (NKRP1A/KLRB1)

    PubMed Central

    Kamishikiryo, Jun; Fukuhara, Hideo; Okabe, Yuki; Kuroki, Kimiko; Maenaka, Katsumi

    2011-01-01

    Human Th17 cells express high levels of CD161, a member of the killer cell lectin-like receptor (KLR) family (also referred to as NK receptor-P1A (NKRP1A) or KLRB1), as a representative marker. CD161 is also expressed on natural killer (NK) cells and NKT cells. Lectin-like transcript 1 (LLT1), another KLR family member, was recently identified as a ligand for CD161. This interaction may play pivotal roles in the immunomodulatory functions of Th17 cells as well as those of NK and NKT cells. However, the molecular basis for the interaction is poorly understood. Here we show that the extracellular domain of CD161 bound directly to LLT1 with a Kd of 48 μm and with the fast kinetics typical of cell-cell recognition receptors. Mutagenesis revealed that the similar membrane-distal β-sheet and loop regions of both CD161 and LLT1 were utilized for the binding, and notably, these regions correspond to the ligand-binding sites for major histocompatibility complex (MHC)-recognizing KLRs. Furthermore, we found a pair of detrimental mutations for both molecules that restored the binding. These results reveal a new template model for the recognition mode between the KLR family members and provide insights into the molecular mechanism underlying Th17/NK/NKT-mediated immune responses. PMID:21572041

  13. Rational design, synthesis, and application of a new receptor for the molecular recognition of tricarboxylate salts in aqueous media.

    PubMed

    Frontera, Antonio; Morey, Jeroni; Oliver, Antonia; Piña, M Neus; Quiñonero, David; Costa, Antoni; Ballester, Pablo; Deyà, Pere M; Anslyn, Eric V

    2006-09-15

    A rational design of a tripodal receptor for the molecular recognition of tricarboxylate salts in aqueous media, based on squaramide, has been performed using high-level DFT calculations (RI-BP86/SVP level of theory) in solution using the COSMO treatment, including some preliminary ab initio calculations at the higher RI-MP2/TZVP level of theory, comparing the ability of squaramide to bind carboxylate salts with two widely used guanidinium salts. The tripodal receptor has been synthesized using a new methodology that has been recently reported by some of us, and its capability of recognizing several mono-, di-, and tricarboxylate salts has been studied experimentally by means of microcalorimetry experiments in a very high competitive media, H(2)O:EtOH 1:3. These experiments give enthalpic and entropic data, which are unfortunately scarce in the literature of molecular recognition of anions. Finally, a fluorimetric ensemble of the receptor with fluorescein has been found to be useful for the fluorimetric determination of zinc citrate in a commercial toothpaste using competition assays. PMID:16958511

  14. Dynamic chemistry of anion recognition

    SciTech Connect

    Custelcean, Radu

    2012-01-01

    In the past 40 years, anion recognition by synthetic receptors has grown into a rich and vibrant research topic, developing into a distinct branch of Supramolecular Chemistry. Traditional anion receptors comprise organic scaffolds functionalized with complementary binding groups that are assembled by multistep organic synthesis. Recently, a new approach to anion receptors has emerged, in which the host is dynamically self-assembled in the presence of the anionic guest, via reversible bond formation between functional building units. While coordination bonds were initially employed for the self-assembly of the anion hosts, more recent studies demonstrated that reversible covalent bonds can serve the same purpose. In both cases, due to their labile connections, the molecular constituents have the ability to assemble, dissociate, and recombine continuously, thereby creating a dynamic combinatorial library (DCL) of receptors. The anionic guests, through specific molecular recognition, may then amplify (express) the formation of a particular structure among all possible combinations (real or virtual) by shifting the equilibria involved towards the most optimal receptor. This approach is not limited to solution self-assembly, but is equally applicable to crystallization, where the fittest anion-binding crystal may be selected. Finally, the pros and cons of employing dynamic combinatorial chemistry (DCC) vs molecular design for developing anion receptors, and the implications of both approaches to selective anion separations, will be discussed.

  15. Application of pattern recognition in molecular spectroscopy: Automatic line search in high-resolution spectra

    NASA Astrophysics Data System (ADS)

    Bykov, A. D.; Pshenichnikov, A. M.; Sinitsa, L. N.; Shcherbakov, A. P.

    2004-07-01

    An expert system has been developed for the initial analysis of a recorded spectrum, namely, for the line search and the determination of line positions and intensities. The expert system is based on pattern recognition algorithms. Object recognition learning allows the system to achieve the needed flexibility and automatically detect groups of overlapping lines, whose profiles should be fit together. Gauss, Lorentz, and Voigt profiles are used as model profiles to which spectral lines are fit. The expert system was applied to processing of the Fourier transform spectrum of the D2O molecule in the region 3200-4200 cm-1, and it detected 4670 lines in the spectrum, which consisted of 439000 dots. No one experimentally observed line exceeding the noise level was missed.

  16. The preparation of magnetic molecularly imprinted nanoparticles for the recognition of bovine hemoglobin.

    PubMed

    Zhang, Min; Wang, Yuzhi; Jia, Xiaoping; He, Meizhi; Xu, Minli; Yang, Shan; Zhang, Cenjin

    2014-03-01

    The protein imprinted technique combining surface imprinting and nano-sized supports materials is an attractive strategy for protein recognition and rapid separation. In this work, we imprinted bovine hemoglobin (BHb) on magnetic nanoparticles. With itaconic acid (IA) and acrylamide (AAm) as the monomers, the experiment was carried out in aqueous media via surface-imprinting technique. The effects of initial concentration and adsorption time over the adsorption capacity of both imprinted and non-imprinted nanoparticles were analyzed. The maximum adsorption capability of imprinted nanoparticles was found to be 77.6 mg g(-1), which was 3.1-4.3 times higher than that of the non-imprinted nanoparticles prepared at the same conditions. This resulted in the successful formation of imprinting cavities. Moreover, in selective adsorption experiment and competitive batch rebinding test, imprinted nanoparticles exhibited a high specific recognition of the template protein over the non-imprinted protein. PMID:24468385

  17. Polymer-based separations: Synthesis and application of polymers for ionic and molecular recognition

    SciTech Connect

    Alexandratos, S.D.

    1992-01-01

    Polymer-based separations have utilized resins such as sulfonic, acrylic, and iminodiacetic acid resins and the XAD series. Selective polymeric reagents for reaction with a targeted metal ion were synthesized as polymers with two different types of functional groups, each operating on the ions through a different mechanism. There are 3 classes of DMBPs (dual mechanism bifunctional polymers). Research during this period dealing with metal ion recognition focused on two of these classes (reduction of metal ions to metal; selective complexation).

  18. Line-Based Object Recognition using Hausdorff Distance: From Range Images to Molecular Secondary Structure

    SciTech Connect

    Guerra, C; Pascucci, V

    2004-12-13

    Object recognition algorithms are fundamental tools in automatic matching of geometric shapes within a background scene. Many approaches have been proposed in the past to solve the object recognition problem. Two of the key aspects that distinguish them in terms of their practical usability are: (i) the type of input model description and (ii) the comparison criteria used. In this paper we introduce a novel scheme for 3D object recognition based on line segment representation of the input shapes and comparison using the Hausdor distance. This choice of model representation provides the flexibility to apply the scheme in different application areas. We define several variants of the Hausdor distance to compare the models within the framework of well defined metric spaces. We present a matching algorithm that efficiently finds a pattern in a 3D scene. The algorithm approximates a minimization procedure of the Hausdor distance. The output error due to the approximation is guaranteed to be within a known constant bound. Practical results are presented for two classes of objects: (i) polyhedral shapes extracted from segmented range images and (ii) secondary structures of large molecules. In both cases the use of our approximate algorithm allows to match correctly the pattern in the background while achieving the efficiency necessary for practical use of the scheme. In particular the performance is improved substantially with minor degradation of the quality of the matching.

  19. Molecular determinants of prokaryotic voltage-gated sodium channels for recognition of local anesthetics.

    PubMed

    Shimomura, Takushi; Irie, Katsumasa; Fujiyoshi, Yoshinori

    2016-08-01

    Local anesthetics (LAs) inhibit mammalian voltage-gated Na(+) channels (Navs) and are thus clinically important. LAs also inhibit prokaryotic Navs (BacNavs), which have a simpler structure than mammalian Navs. To elucidate the detailed mechanisms of LA inhibition to BacNavs, we used NavBh, a BacNav from Bacillus halodurans, to analyze the interactions of several LAs and quaternary ammoniums (QAs). Based on the chemical similarity of QA with the tertiary-alkylamine (TAA) group of LAs, QAs were used to determine the residues required for the recognition of TAA by NavBh. We confirmed that two residues, Thr220 and Phe227, are important for LA binding; a methyl group of Thr220 is important for recognizing both QAs and LAs, whereas Phe227 is involved in holding blockers at the binding site. In addition, we found that NavBh holds blockers in a closed state, consistent with the large inner cavity observed in the crystal structures of BacNavs. These findings reveal the inhibition mechanism of LAs in NavBh, where the methyl group of Thr220 provides the main receptor site for the TAA group and the bulky phenyl group of Phe227 holds the blockers inside the large inner cavity. These two residues correspond to the two LA recognition residues in mammalian Navs, which suggests the relevance of the LA recognition between BacNavs and mammalian Navs. PMID:27273848

  20. The role of conformational selection in the molecular recognition of the wild type and mutants XPA67-80 peptides by ERCC1.

    PubMed

    Fadda, Elisa

    2015-07-01

    Molecular recognition is a fundamental step in the coordination of biomolecular pathways. Understanding how recognition and binding occur between highly flexible protein domains is a complex task. The conformational selection theory provides an elegant rationalization of the recognition mechanism, especially valid in cases when unstructured protein regions are involved. The recognition of a poorly structured peptide, namely XPA67-80 , by its target receptor ERCC1, falls in this challenging study category. The microsecond molecular dynamics (MD) simulations, discussed in this work, show that the conformational propensity of the wild type XPA67-80 peptide in solution supports conformational selection as the key mechanism driving its molecular recognition by ERCC1. Moreover, all the mutations of the XPA67-80 peptide studied here cause a significant increase of its conformational disorder, relative to the wild type. Comparison to experimental data suggests that the loss of the recognized structural motifs at the microscopic time scale can contribute to the critical decrease in binding observed for one of the mutants, further substantiating the key role of conformational selection in recognition. Ultimately, because of the high sequence identity and analogy in binding, it is conceivable that the conclusions of this study on the XPA67-80 peptide also apply to the ERCC1-binding domain of the XPA protein. PMID:25973722

  1. A molecular switch in the scaffold NHERF1 enables misfolded CFTR to evade the peripheral quality control checkpoint.

    PubMed

    Loureiro, Cláudia A; Matos, Ana Margarida; Dias-Alves, Ângela; Pereira, Joana F; Uliyakina, Inna; Barros, Patrícia; Amaral, Margarida D; Matos, Paulo

    2015-05-19

    The peripheral protein quality control (PPQC) checkpoint removes improperly folded proteins from the plasma membrane through a mechanism involving the E3 ubiquitin ligase CHIP (carboxyl terminus of Hsc70 interacting protein). PPQC limits the efficacy of some cystic fibrosis (CF) drugs, such as VX-809, that improve trafficking to the plasma membrane of misfolded mutants of the CF transmembrane conductance regulator (CFTR), including F508del-CFTR, which retains partial functionality. We investigated the PPQC checkpoint in lung epithelial cells with F508del-CFTR that were exposed to VX-809. The conformation of the scaffold protein NHERF1 (Na(+)/H(+) exchange regulatory factor 1) determined whether the PPQC recognized "rescued" F508del-CFTR (the portion that reached the cell surface in VX-809-treated cells). Activation of the cytoskeletal regulator Rac1 promoted an interaction between the actin-binding adaptor protein ezrin and NHERF1, triggering exposure of the second PDZ domain of NHERF1, which interacted with rescued F508del-CFTR. Because binding of F508del-CFTR to the second PDZ of NHERF1 precluded the recruitment of CHIP, the coexposure of airway cells to Rac1 activator nearly tripled the efficacy of VX-809. Interference with the NHERF1-ezrin interaction prevented the increase of efficacy of VX-809 by Rac1 activation, but the actin-binding domain of ezrin was not required for the increase in efficacy. Thus, rather than mainly directing anchoring of F508del-CFTR to the actin cytoskeleton, induction of ezrin activation by Rac1 signaling triggered a conformational change in NHERF1, which was then able to bind and stabilize misfolded CFTR at the plasma membrane. These insights into the cell surface stabilization of CFTR provide new targets to improve treatment of CF. PMID:25990958

  2. Quantification of the transferability of a designed protein specificity switch reveals extensive epistasis in molecular recognition

    SciTech Connect

    Melero, Cristina; Ollikainen, Noah; Harwood, Ian; Karpiak, Joel; Kortemme, Tanja

    2014-10-13

    Re-engineering protein–protein recognition is an important route to dissecting and controlling complex interaction networks. Experimental approaches have used the strategy of “second-site suppressors,” where a functional interaction is inferred between two proteins if a mutation in one protein can be compensated by a mutation in the second. Mimicking this strategy, computational design has been applied successfully to change protein recognition specificity by predicting such sets of compensatory mutations in protein–protein interfaces. To extend this approach, it would be advantageous to be able to “transplant” existing engineered and experimentally validated specificity changes to other homologous protein–protein complexes. Here, we test this strategy by designing a pair of mutations that modulates peptide recognition specificity in the Syntrophin PDZ domain, confirming the designed interaction biochemically and structurally, and then transplanting the mutations into the context of five related PDZ domain–peptide complexes. We find a wide range of energetic effects of identical mutations in structurally similar positions, revealing a dramatic context dependence (epistasis) of designed mutations in homologous protein–protein interactions. To better understand the structural basis of this context dependence, we apply a structure-based computational model that recapitulates these energetic effects and we use this model to make and validate forward predictions. The context dependence of these mutations is captured by computational predictions, our results both highlight the considerable difficulties in designing protein–protein interactions and provide challenging benchmark cases for the development of improved protein modeling and design methods that accurately account for the context.

  3. Quantification of the transferability of a designed protein specificity switch reveals extensive epistasis in molecular recognition

    DOE PAGESBeta

    Melero, Cristina; Ollikainen, Noah; Harwood, Ian; Karpiak, Joel; Kortemme, Tanja

    2014-10-13

    Re-engineering protein–protein recognition is an important route to dissecting and controlling complex interaction networks. Experimental approaches have used the strategy of “second-site suppressors,” where a functional interaction is inferred between two proteins if a mutation in one protein can be compensated by a mutation in the second. Mimicking this strategy, computational design has been applied successfully to change protein recognition specificity by predicting such sets of compensatory mutations in protein–protein interfaces. To extend this approach, it would be advantageous to be able to “transplant” existing engineered and experimentally validated specificity changes to other homologous protein–protein complexes. Here, we test thismore » strategy by designing a pair of mutations that modulates peptide recognition specificity in the Syntrophin PDZ domain, confirming the designed interaction biochemically and structurally, and then transplanting the mutations into the context of five related PDZ domain–peptide complexes. We find a wide range of energetic effects of identical mutations in structurally similar positions, revealing a dramatic context dependence (epistasis) of designed mutations in homologous protein–protein interactions. To better understand the structural basis of this context dependence, we apply a structure-based computational model that recapitulates these energetic effects and we use this model to make and validate forward predictions. The context dependence of these mutations is captured by computational predictions, our results both highlight the considerable difficulties in designing protein–protein interactions and provide challenging benchmark cases for the development of improved protein modeling and design methods that accurately account for the context.« less

  4. STUDIES OF RELATIONSHIPS BETWEEN MOLECULAR STRUCTURE AND BIOLOGICAL ACTIVITY BY PATTERN RECOGNITION METHODS

    EPA Science Inventory

    The attempt to rationalize the connections between the molecular structures of organic compounds and their biological activities comprises the field of structure-activity relations (SAR) studies. Correlations between structure and activity are important for the understanding and ...

  5. A molecular mechanism realizing sequence-specific recognition of nucleic acids by TDP-43

    PubMed Central

    Furukawa, Yoshiaki; Suzuki, Yoh; Fukuoka, Mami; Nagasawa, Kenichi; Nakagome, Kenta; Shimizu, Hideaki; Mukaiyama, Atsushi; Akiyama, Shuji

    2016-01-01

    TAR DNA-binding protein 43 (TDP-43) is a DNA/RNA-binding protein containing two consecutive RNA recognition motifs (RRM1 and RRM2) in tandem. Functional abnormality of TDP-43 has been proposed to cause neurodegeneration, but it remains obscure how the physiological functions of this protein are regulated. Here, we show distinct roles of RRM1 and RRM2 in the sequence-specific substrate recognition of TDP-43. RRM1 was found to bind a wide spectrum of ssDNA sequences, while no binding was observed between RRM2 and ssDNA. When two RRMs are fused in tandem as in native TDP-43, the fused construct almost exclusively binds ssDNA with a TG-repeat sequence. In contrast, such sequence-specificity was not observed in a simple mixture of RRM1 and RRM2. We thus propose that the spatial arrangement of multiple RRMs in DNA/RNA binding proteins provides steric effects on the substrate-binding site and thereby controls the specificity of its substrate nucleotide sequences. PMID:26838063

  6. A metal-catalyzed enyne-cyclization step for the synthesis of bi- and tricyclic scaffolds amenable to molecular library production.

    PubMed

    Wu, Peng; Petersen, Michael Åxman; Cohrt, A Emil; Petersen, Rico; Morgentin, Rémy; Lemoine, Hugues; Roche, Carine; Willaume, Anthony; Clausen, Mads H; Nielsen, Thomas E

    2016-08-01

    A facile metal-catalyzed diversification step for the synthesis of novel bi- and tricyclic scaffolds from enyne substrates is reported in this study. From a single starting material, topologically diverse scaffolds for library synthesis can be generated and decorated in a few steps. The methodology was used to produce a library of 490 compounds within the European Lead Factory (ELF) Consortium. PMID:27356738

  7. Exploring the ligand recognition properties of the human vasopressin V1a receptor using QSAR and molecular modeling studies.

    PubMed

    Contreras-Romo, Martha C; Martínez-Archundia, Marlet; Deeb, Omar; Slusarz, Magdalena J; Ramírez-Salinas, Gema; Garduño-Juárez, Ramón; Quintanar-Stephano, Andrés; Ramírez-Galicia, Guillermo; Correa-Basurto, José

    2014-02-01

    Vaptans are compounds that act as non-peptide vasopressin receptor antagonists. These compounds have diverse chemical structures. In this study, we used a combined approach of protein folding, molecular dynamics simulations, docking, and quantitative structure-activity relationship (QSAR) to elucidate the detailed interaction of the vasopressin receptor V1a (V1aR) with some of its blockers (134). QSAR studies were performed using MLR analysis and were gathered into one group to perform an artificial neural network (ANN) analysis. For each molecule, 1481 molecular descriptors were calculated. Additionally, 15 quantum chemical descriptors were calculated. The final equation was developed by choosing the optimal combination of descriptors after removing the outliers. Molecular modeling enabled us to obtain a reliable tridimensional model of V1aR. The docking results indicated that the great majority of ligands reach the binding site under π-π, π-cation, and hydrophobic interactions. The QSAR studies demonstrated that the heteroatoms N and O are important for ligand recognition, which could explain the structural diversity of ligands that reach V1aR. PMID:24010681

  8. Superhydrophilic molecularly imprinted polymers based on a water-soluble functional monomer for the recognition of gastrodin in water media.

    PubMed

    Ji, Wenhua; Zhang, Mingming; Wang, Daijie; Wang, Xiao; Liu, Jianhua; Huang, Luqi

    2015-12-18

    In this study, the first successfully developed superhydrophilic molecularly imprinted polymers (MIPs) for gastrodin recognition have been described. MIPs were prepared via the bulk polymerization process in an aqueous solution using alkenyl glycosides glucose (AGG) as the water-soluble functional monomer. The non-imprinted polymers (NIPs) were also synthesized using the same method without the use of the template. The dynamic water contact angles and photographs of the dispersion properties confirmed that the molecularly imprinted polymers displayed excellent superhydrophilicity. The results demonstrated that the MIPs exhibited high selectivity and an excellent imprinting effect. A molecularly imprinted solid phase extraction (MISPE) method was established. Optimization of various parameters affecting MISPE was investigated. Under the optimized conditions, a wide linear range (0.001-100.0μgmL(-1)) and low limits of detection (LOD) and quantification (LOQ) (0.03 and 0.09ngmL(-1), respectively) were achieved. When compared with the NIPs, higher recoveries (90.5% to 97.6%) of gastrodin with lower relative standard deviations values (below 6.4%) using high performance liquid chromatography were obtained at three spiked levels in three blank samples. These results demonstrated one efficient, highly selective and environmentally-friendly MISPE technique with excellent reproducibility for the purification and pre-concentration of gastrodin from an aqueous extract of Gastrodia elata roots. PMID:26627582

  9. The challenges involved in elucidating the molecular basis of sperm-egg recognition in mammals and approaches to overcome them.

    PubMed

    Wright, Gavin J; Bianchi, Enrica

    2016-01-01

    Sexual reproduction is used by many different organisms to create a new generation of genetically distinct progeny. Cells originating from separate sexes or mating types segregate their genetic material into haploid gametes which must then recognize and fuse with each other in a process known as fertilization to form a diploid zygote. Despite the central importance of fertilization, we know remarkably little about the molecular mechanisms that are involved in how gametes recognize each other, particularly in mammals, although the proteins that are displayed on their surfaces are almost certainly involved. This paucity of knowledge is largely due to both the unique biological properties of mammalian gametes (sperm and egg) which make them experimentally difficult to manipulate, and the technical challenges of identifying interactions between membrane-embedded cell surface receptor proteins. In this review, we will discuss our current knowledge of animal gamete recognition, highlighting where important contributions to our understanding were made, why particular model systems were helpful, and why progress in mammals has been particularly challenging. We discuss how the development of mammalian in vitro fertilization and targeted gene disruption in mice were important technological advances that triggered progress. We argue that approaches employed to discover novel interactions between cell surface gamete recognition proteins should account for the unusual biochemical properties of membrane proteins and the typically highly transient nature of their interactions. Finally, we describe how these principles were applied to identify Juno as the egg receptor for sperm Izumo1, an interaction that is essential for mammalian fertilization. PMID:26224538

  10. Effective Use of Molecular Recognition in Gas Sensing: Results from Acoustic Wave and In-Situ FTIR Measurements

    SciTech Connect

    Bodenhofer, K,; Gopel, W.; Hierlemann, A.; Ricco, A.J.

    1998-12-09

    To probe directly the analyte/film interactions that characterize molecular recognition in gas sensors, we recorded changes to the in-situ surface vibrational spectra of specifically fictionalized surface acoustic wave (SAW) devices concurrently with analyte exposure and SAW measurement of the extent of sorption. Fourier-lmnsform infrared external- reflectance spectra (FTIR-ERS) were collected from operating 97-MH2 SAW delay lines during exposure to a range of analytes as they interacted with thin-film coatings previously shown to be selective: cyclodextrins for chiral recognition, Ni-camphorates for Lewis bases such as pyridine and organophosphonates, and phthalocyanines for aromatic compounds. In most cases where specific chemical interactions-metal coordination, "cage" compound inclusion, or z stacking-were expected, analyte dosing caused distinctive changes in the IR spectr~ together with anomalously large SAW sensor responses. In contrast, control experiments involving the physisorption of the same analytes by conventional organic polymers did not cause similar changes in the IR spectra, and the SAW responses were smaller. For a given conventional polymer, the partition coefficients (or SAW sensor signals) roughly followed the analyte fraction of saturation vapor pressure. These SAW/FTIR results support earlier conclusions derived from thickness-shear mode resonator data.

  11. Assessing the potential of atomistic molecular dynamics simulations to probe reversible protein-protein recognition and binding

    PubMed Central

    Abriata, Luciano A.; Dal Peraro, Matteo

    2015-01-01

    Protein-protein recognition and binding are governed by diffusion, noncovalent forces and conformational flexibility, entangled in a way that only molecular dynamics simulations can dissect at high resolution. Here we exploited ubiquitin’s noncovalent dimerization equilibrium to assess the potential of atomistic simulations to reproduce reversible protein-protein binding, by running submicrosecond simulations of systems with multiple copies of the protein at millimolar concentrations. The simulations essentially fail because they lead to aggregates, yet they reproduce some specificity in the binding interfaces as observed in known covalent and noncovalent ubiquitin dimers. Following similar observations in literature we hint at electrostatics and water descriptions as the main liable force field elements, and propose that their optimization should consider observables relevant to multi-protein systems and unfolded proteins. Within limitations, analysis of binding events suggests salient features of protein-protein recognition and binding, to be retested with improved force fields. Among them, that specific configurations of relative direction and orientation seem to trigger fast binding of two molecules, even over 50 Å distances; that conformational selection can take place within surface-to-surface distances of 10 to 40 Å i.e. well before actual intermolecular contact; and that establishment of contacts between molecules further locks their conformations and relative orientations. PMID:26023027

  12. Design, synthesis, and biological evaluation of the combinatorial library with a new spirodiketopiperazine scaffold. Discovery of novel potent and selective low-molecular-weight CCR5 antagonists.

    PubMed

    Habashita, Hiromu; Kokubo, Masaya; Hamano, Shin-ichi; Hamanaka, Nobuyuki; Toda, Masaaki; Shibayama, Shiro; Tada, Hideaki; Sagawa, Kenji; Fukushima, Daikichi; Maeda, Kenji; Mitsuya, Hiroaki

    2006-07-13

    We previously reported the discovery of several spirodiketopiperazine derivatives as potent CCR5 antagonists with anti-HIV activity. Herein, we describe in detail the identification of these lead compounds using a combinatorial chemistry approach. A novel spirodiketopiperazine scaffold was designed on the basis of the concept of the privileged structure of G-protein-coupled receptors (GPCRs). This new framework was obtained in acceptable yield with high purity from the readily prepared isonitrile resin through the Ugi reaction, sequential transformations, and cyclative cleavage. By measuring the inhibitory activity of each compound in the initial library against the intracellular calcium mobilization stimulated by MIP-1alpha, several compounds were found to show modest but selective CCR5 antagonistic activity. After the rapid evaluation of these hit compounds, several single-digit nanomolar, low-molecular-weight CCR5 antagonists that can potently block the infectivity and replication of laboratory and clinical strains of HIV as well as those of highly drug-resistant HIV variants with minimal cytotoxicity have been identified. PMID:16821774

  13. Ion and molecule sensors using molecular recognition in luminescent, conductive polymers. FY 1997 year-end progress report

    SciTech Connect

    Wasielewski, M.R.

    1997-01-01

    'The purpose of this project is to use molecular recognition strategies to develop sensor technology based on luminescent, conductive polymers that contain sites for binding specific molecules or ions in the presence of related molecules or ions. Selective binding of a particular molecule or ion of interest to these polymers will result in a large change in their luminescence and/or conductivity, which can be used to both qualitatively and quantitatively sense the presence of the bound molecules or ions. The main thrusts and accomplishments in the first year of this project involve developing polymer syntheses that yield conjugated polymers to which a wide variety of ligands for metal ion binding can be readily incorporated.'

  14. Competitive fluorescence assay for specific recognition of atrazine by magnetic molecularly imprinted polymer based on Fe3O4-chitosan.

    PubMed

    Liu, Guangyang; Li, Tengfei; Yang, Xin; She, Yongxin; Wang, Miao; Wang, Jing; Zhang, Min; Wang, Shanshan; Jin, Fen; Jin, Maojun; Shao, Hua; Jiang, Zejun; Yu, Hailong

    2016-02-10

    A novel fluorescence sensing strategy for determination of atrazine in tap water involving direct competition between atrazine and 5-(4,6-dichlorotriazinyl) aminofluorescein (5-DTAF), and which exploits magnetic molecularly imprinted polymer (MMIP), has been developed. The MMIP, based on Fe3O4-chitosan nanoparticles, was synthesized to recognize specific binding sites of atrazine. The recognition capability and selectivity of the MMIP for atrazine and other triazine herbicides was investigated. Under optimal conditions, the competitive reaction between 5-DTAF and atrazine was performed to permit quantitation. Fluorescence intensity changes at 515 nm was linearly related to the logarithm of the atrazine concentration for the range 2.32-185.4 μM. The detection limit for atrazine was 0.86μM (S/N=3) and recoveries were 77.6-115% in spiked tap water samples. PMID:26686107

  15. Helicase-Dependent Isothermal Amplification of DNA and RNA by Using Self-Avoiding Molecular Recognition Systems.

    PubMed

    Yang, Zunyi; McLendon, Chris; Hutter, Daniel; Bradley, Kevin M; Hoshika, Shuichi; Frye, Carole B; Benner, Steven A

    2015-06-15

    Assays that detect DNA or RNA (xNA) are highly sensitive, as small amounts of xNA can be amplified by PCR. Unfortunately, PCR is inconvenient in low-resource environments, and requires equipment and power that might not be available in these environments. Isothermal procedures, which avoid thermal cycling, are often confounded by primer dimers, off-target priming, and other artifacts. Here, we show how a "self avoiding molecular recognition system" (SAMRS) eliminates these artifacts and gives clean amplicons in a helicase-dependent isothermal amplification (SAMRS-HDA). We also show that incorporating SAMRS into the 3'-ends of primers facilitates the design and screening of primers for HDA assays. Finally, we show that SAMRS-HDA can be twofold multiplexed, difficult to achieve with HDA using standard primers. Thus, SAMRS-HDA is a more versatile approach than standard HDA, with a broader applicability for xNA-targeted diagnostics and research. PMID:25953623

  16. Molecular recognition of curcumin (Indian Ayurvedic medicine) by the supramolecular probe, p-t-butyl calix(8)arene

    NASA Astrophysics Data System (ADS)

    Meenakshi, C.; Jayabal, P.; Ramakrishnan, V.

    2014-06-01

    The thermodynamic property of the host-guest complexes formed between the curcumin, component of Indian Ayurvedic medicine turmeric, a drug molecule, with the supra molecule, p-t-butyl calix(8)arene was studied. p-t-Butyl calix(8)arene has been used as a host molecule and curcumin as a guest molecule. Optical absorption spectral studies were carried out to investigate the molecular recognition properties of p-t-butyl calix(8)arene with curcumin. The stochiometry of the host-guest complexes formed and the binding constant were determined. An interesting 1:1 and 4:1 stochiometric host-guest complexes were formed. Job's continuous method of variation and Benesi-Hildebrand expression were used for the determination of binding constant and the stochiometry of the host-guest complex formed.

  17. New biomedical devices with selective peptide recognition properties. Part 1: Characterization and cytotoxicity of molecularly imprinted polymers

    PubMed Central

    Rechichi, A; Cristallini, C; Vitale, U; Ciardelli, G; Barbani, N; Vozzi, G; Giusti, P

    2007-01-01

    Abstract Molecular imprinting is a technique for the synthesis of polymers capable to bind target molecules selectively. The imprinting of large proteins, such as cell adhesion proteins or cell receptors, opens the way to important and innovative biomedical applications. However, such molecules can incur into important conformational changes during the preparation of the imprinted polymer impairing the specificity of the recognition cavities. The “epitope approach” can overcome this limit by adopting, as template, a short peptide sequence representative of an accessible fragment of a larger protein. The resulting imprinted polymer can recognize both the template and the whole molecule thanks to the specific cavities for the epitope. In this work two molecularly imprinted polymer formulations (a macroporous monolith and nanospheres) were obtained using the protected peptide Z-Thr-Ala-Ala-OMe, as template, and Z-Thr-Ile-Leu-OMe, as analogue for the selectivity evaluation, methacrylic acid, as functional monomer, and trimethylolpropane trimethacrylate and pentaerythritol triacrylate (PETRA), as cross-linkers. Polymers were synthesized by precipitation polymerization and characterized by standard techniques. Polymerization and rebinding solutions were analyzed by high performance liquid chromatography. The highly cross-linked polymers retained about 70% of the total template amount, against (20% for the less cross-linked ones). The extracted template amount and the rebinding capacity decreased with the cross-linking degree, while the selectivity showed the opposite behaviour. The PETRA cross-linked polymers showed the best recognition (MIP 2−, α= 1.71) and selectivity (MIP 2+, α′= 5.58) capabilities. The cytotoxicity tests showed normal adhesion and proliferation of fibroblasts cultured in the medium that was put in contact with the imprinted polymers. PMID:18205706

  18. Density functional theory based study of molecular interactions, recognition, engineering, and quantum transport in π molecular systems.

    PubMed

    Cho, Yeonchoo; Cho, Woo Jong; Youn, Il Seung; Lee, Geunsik; Singh, N Jiten; Kim, Kwang S

    2014-11-18

    CONSPECTUS: In chemical and biological systems, various interactions that govern the chemical and physical properties of molecules, assembling phenomena, and electronic transport properties compete and control the microscopic structure of materials. The well-controlled manipulation of each component can allow researchers to design receptors or sensors, new molecular architectures, structures with novel morphology, and functional molecules or devices. In this Account, we describe the structures and electronic and spintronic properties of π-molecular systems that are important for controlling the architecture of a variety of carbon-based systems. Although DFT is an important tool for describing molecular interactions, the inability of DFT to accurately represent dispersion interactions has made it difficult to properly describe π-interactions. However, the recently developed dispersion corrections for DFT have allowed us to include these dispersion interactions cost-effectively. We have investigated noncovalent interactions of various π-systems including aromatic-π, aliphatic-π, and non-π systems based on dispersion-corrected DFT (DFT-D). In addition, we have addressed the validity of DFT-D compared with the complete basis set (CBS) limit values of coupled cluster theory with single, double, and perturbative triple excitations [CCSD(T)] and Møller-Plesset second order perturbation theory (MP2). The DFT-D methods are still unable to predict the correct ordering in binding energies within the benzene dimer and the cyclohexane dimer. Nevertheless, the overall DFT-D predicted binding energies are in reasonable agreement with the CCSD(T) results. In most cases, results using the B97-D3 method closely reproduce the CCSD(T) results with the optimized energy-fitting parameters. On the other hand, vdW-DF2 and PBE0-TS methods estimate the dispersion energies from the calculated electron density. In these approximations, the interaction energies around the equilibrium

  19. Cardiac-Oxidized Antigens Are Targets of Immune Recognition by Antibodies and Potential Molecular Determinants in Chagas Disease Pathogenesis

    PubMed Central

    Dhiman, Monisha; Zago, Maria Paola; Nunez, Sonia; Amoroso, Alejandro; Rementeria, Hugo; Dousset, Pierre; Burgos, Federico Nunez; Garg, Nisha Jain

    2012-01-01

    conclude that ROS-induced, cardiac-oxidized antigens are targets of immune recognition by antibodies and molecular determinants for pathogenesis during Chagas disease. PMID:22238578

  20. Molecular Mechanisms of Substrate Recognition and Specificity of New Delhi Metallo-β-Lactamase

    PubMed Central

    Chiou, Jiachi; Leung, Thomas Yun-Chung

    2014-01-01

    Carbapenems are one of the last lines of defense for Gram-negative pathogens, such as members of the Enterobacteriaceae. Despite the fact that most carbapenems are resistant to extended-spectrum β-lactamase (ESBL), emerging metallo-β-lactamases (MBLs), including New Delhi metallo-β-lactamase 1 (NDM-1), that can hydrolyze carbapenems have become prevalent and are frequently associated with the so-called “superbugs,” for which treatments are extremely limited. Crystallographic study sheds light on the modes of antibiotic binding to NDM-1, yet the mechanisms governing substrate recognition and specificity are largely unclear. This study provides a connection between crystallographic study and the functional significance of NDM-1, with an emphasis on the substrate specificity and catalysis of various β-lactams. L1 loop residues L59, V67, and W87 were important for the activity of NDM-1, most likely through maintaining the partial folding of the L1 loop or active site conformation through hydrophobic interaction with the R groups of β-lactams or the β-lactam ring. Substitution of alanine for L59 showed greater reduction of MICs to ampicillin and selected cephalosporins, whereas substitutions of alanine for V67 had more impact on the MICs of carbapenems. K224 and N233 on the L3 loop played important roles in the recognition of substrate and contributed to substrate hydrolysis. These data together with the structure comparison of the B1 and B2 subclasses of MBLs revealed that the broad substrate specificity of NDM-1 could be due to the ability of its wide active site cavity to accommodate a wide range of β-lactams. This study provides insights into the development of efficient inhibitors for NDM-1 and offers an efficient tactic with which to study the substrate specificities of other β-lactamases. PMID:24982075

  1. Molecular recognition in gas sensing: Results from acoustic wave and in-situ FTIR measurements

    SciTech Connect

    Hierlemann, A.; Ricco, A.J.; Bodenhoefer, K.; Goepel, W.

    1998-06-01

    Surface acoustic wave (SAW) measurements were combined with direct, in-situ molecular spectroscopy to understand the interactions of surface-confined sensing films with gas-phase analytes. This was accomplished by collecting Fourier-transform infrared external-reflectance spectra (FTIR-ERS) on operating SAW devices during dosing of their specifically coated surfaces with key analytes.

  2. Application of Machine Learning tools to recognition of molecular patterns in STM images

    NASA Astrophysics Data System (ADS)

    Maksov, Artem; Ziatdinov, Maxim; Fujii, Shintaro; Kiguchi, Manabu; Higashibayashi, Shuhei; Sakurai, Hidehiro; Kalinin, Sergei; Sumpter, Bobby

    The ability to utilize individual molecules and molecular assemblies as data storage elements has motivated scientist for years, concurrent with the continuous effort to shrink a size of data storage devices in microelectronics industry. One of the critical issues in this effort lies in being able to identify individual molecular assembly units (patterns), on a large scale in an automated fashion of complete information extraction. Here we present a novel method of applying machine learning techniques for extraction of positional and rotational information from scanning tunneling microscopy (STM) images of π-bowl sumanene molecules on gold. We use Markov Random Field (MRF) model to decode the polar rotational states for each molecule in a large scale STM image of molecular film. We further develop an algorithm that uses a convolutional Neural Network combined with MRF and input from density functional theory to classify molecules into different azimuthal rotational classes. Our results demonstrate that a molecular film is partitioned into distinctive azimuthal rotational domains consisting typically of 20-30 molecules. In each domain, the ``bowl-down'' molecules are generally surrounded by six nearest neighbor molecules in ``bowl-up'' configuration, and the resultant overall structure form a periodic lattice of rotational and polar states within each domain. Research was supported by the US Department of Energy.

  3. Electron sharing and anion-π recognition in molecular triangular prisms.

    PubMed

    Schneebeli, Severin T; Frasconi, Marco; Liu, Zhichang; Wu, Yilei; Gardner, Daniel M; Strutt, Nathan L; Cheng, Chuyang; Carmieli, Raanan; Wasielewski, Michael R; Stoddart, J Fraser

    2013-12-01

    Stacking on a full belly: Triangular molecular prisms display electron sharing among their triangularly arranged naphthalenediimide (NDI) redox centers. Their electron-deficient cavities encapsulate linear triiodide anions, leading to the formation of supramolecular helices in the solid state. Chirality transfer is observed from the six chiral centers of the filled prisms to the single-handed helices. PMID:24227594

  4. A phosphonic acid appended naphthalene diimide motif for self-assembly into tunable nanostructures through molecular recognition with arginine in water.

    PubMed

    Nandre, Kamalakar P; Bhosale, Sheshanath V; Rama Krishna, K V S; Gupta, Akhil; Bhosale, Sidhanath V

    2013-06-18

    A naphthalene diimide motif bearing phosphonic acid functionalities has been found to be self-assembled with L- and D-arginine through chirality induced molecular recognitions and leads to the formation of micrometre long nanobelts and spherical aggregates at pH 9 in water, respectively. PMID:23589823

  5. The molecular recognition of dipeptide by oligoglycyl head group of amphiphile: a quantum chemical study.

    PubMed

    Thirumoorthy, Krishnan; Soni, Kiran; Nandi, Nilashis

    2009-01-01

    In the present work, we presented an analysis of the unusual recognition specificity exhibited by marked difference in the binding behavior of dipeptide with amphiphilic head group when subtle relative change of N-terminal and C-terminal of the dipeptide are made. Recently, in a series of detailed experiments, binding of aqueous dipeptides, GlyX and X/Gly (X = Leu, Phe, Pro, Ala; X/ = Leu, Phe) with dialkyl oligoglycyl amphiphiles is studied [X. Cha, K. Ariga, M. Onda, and T. Kunitake, J. Am. Chem. Soc. 117, 11833 (1995)]. It is observed that GlyX are specifically bound to 2C18BGly2NH2 while X/Gly are insignificantly bound. We first studied the conformational energy variation of GlyPhe, PheGly and model of 2C18BGly2NH2 amphiphile using semi-empirical and ab-initio methods in vacuum. Using the individual energy optimized monomer structure of amphiphile and peptide, we studied the binding energy of optimized GlyPhe: amphiphile pair and PheGly: amphiphile pair structures at 1:1 and 1:2 ratio at the same level of theory using a population of structures. Binding of GlyPhe is favorable over the binding of PheGly at various levels of theory (semi-empirical and ab-initio). It is noted that the hydrogen bonding pattern in the GlyPhe binding is more effective than that in the PheGly binding. In the population of low energy structures, PheGly: amphiphile structures have more exposed area around the hydrophobic Phe group than the GlyPhe: amphiphile structures. Relatively more PheGly: amphiphile structures have intermolecular orientation unsuitable to contribute to the population of head group structures relevant in aqueous interface. Summarizing, significantly better binding capacity of GlyPhe over the PheGly with amphiphile, is due to the difference in hydrogen bonding interaction pattern, hydrophobic effect and possible orientations of the amphiphile and peptide at interface, relevant to the condensed phase monolayer structure. All the three factors cooperatively lead to

  6. 2-Phenylpyrazolo[4,3-d]pyrimidin-7-one as a new scaffold to obtain potent and selective human A3 adenosine receptor antagonists: new insights into the receptor-antagonist recognition.

    PubMed

    Lenzi, Ombretta; Colotta, Vittoria; Catarzi, Daniela; Varano, Flavia; Poli, Daniela; Filacchioni, Guido; Varani, Katia; Vincenzi, Fabrizio; Borea, Pier Andrea; Paoletta, Silvia; Morizzo, Erika; Moro, Stefano

    2009-12-10

    A molecular simplification approach of previously reported 2-arylpyrazolo[3,4-c]quinolin-4-ones was applied to design 2-arylpyrazolo[4,3-d]pyrimidin-7-one derivatives as new human A(3) adenosine receptor antagonists. Substituents with different lipophilicity and steric hindrance were introduced at the 5-position of the bicyclic scaffold (R(5) = H, Me, Et, Ph, CH(2)Ph) and on the 2-phenyl ring (OMe, Me). Most of the synthesized derivatives were highly potent hA(3) adenosine receptor antagonists, the best being the 2-(4-methoxyphenyl)pyrazolo[4,3-d]pyrimidin-7-one (K(i) = 1.2 nM). The new compounds were also highly selective, being completely devoid of affinity toward hA(1), hA(2A), and hA(2B) adenosine receptors. On the basis of the recently published human A(2A) receptor crystallographic information, we propose a novel receptor-driven hypothesis to explain both A(3) AR affinity and A(3) versus A(2A) selectivity profiles of these new antagonists. PMID:19743865

  7. Preparation and Application of Novel Magnetic Molecularly Imprinted Composites for Recognition of Sulfadimethoxine in Feed Samples.

    PubMed

    Feng, Min; Li, Hengye; Zhang, Lin; Zhang, Jingyou; Dai, Jianping; Wang, Xiaojin; Zhang, Lingli; Wei, Yunji

    2016-01-01

    Novel magnetic molecularly imprinted composites were prepared through a facile method using sulfadimethoxine (SDM) as template. The inorganic magnetic nanoparticles were linked with the organic molecularly imprinted polymer (MIP) through irreversibly covalent bond. So, the resulted composites showed excellent stability and reusability under acidic elution conditions. The magnetic MIP composites showed good selectivity, high binding capacity and excellent kinetics toward SDM. Adopting the magnetic MIP composites as extraction material, an off-line magnetic solid-phase extraction (SPE)/high performance liquid chromatography (HPLC) method was established. The calibration curve was linear in the range of 0.05 - 15 mg kg(-1) (r(2) = 0.9976). The LOD and LOQ were 0.016 and 0.052 mg kg(-1), respectively, while the recoveries were in the range of 89.3 - 107.0%. These novel magnetic MIP composites may become a powerful tool for the extraction of template from complex samples with good efficiency. PMID:27169650

  8. Large scale affinity calculations of cyclodextrin host-guest complexes: Understanding the role of reorganization in the molecular recognition process

    PubMed Central

    Wickstrom, Lauren; He, Peng; Gallicchio, Emilio; Levy, Ronald M.

    2013-01-01

    Host-guest inclusion complexes are useful models for understanding the structural and energetic aspects of molecular recognition. Due to their small size relative to much larger protein-ligand complexes, converged results can be obtained rapidly for these systems thus offering the opportunity to more reliably study fundamental aspects of the thermodynamics of binding. In this work, we have performed a large scale binding affinity survey of 57 β-cyclodextrin (CD) host guest systems using the binding energy distribution analysis method (BEDAM) with implicit solvation (OPLS-AA/AGBNP2). Converged estimates of the standard binding free energies are obtained for these systems by employing techniques such as parallel Hamitionian replica exchange molecular dynamics, conformational reservoirs and multistate free energy estimators. Good agreement with experimental measurements is obtained in terms of both numerical accuracy and affinity rankings. Overall, average effective binding energies reproduce affinity rank ordering better than the calculated binding affinities, even though calculated binding free energies, which account for effects such as conformational strain and entropy loss upon binding, provide lower root mean square errors when compared to measurements. Interestingly, we find that binding free energies are superior rank order predictors for a large subset containing the most flexible guests. The results indicate that, while challenging, accurate modeling of reorganization effects can lead to ligand design models of superior predictive power for rank ordering relative to models based only on ligand-receptor interaction energies. PMID:25147485

  9. Molecular mechanism of ligand recognition by membrane transport protein, Mhp1

    PubMed Central

    Simmons, Katie J; Jackson, Scott M; Brueckner, Florian; Patching, Simon G; Beckstein, Oliver; Ivanova, Ekaterina; Geng, Tian; Weyand, Simone; Drew, David; Lanigan, Joseph; Sharples, David J; Sansom, Mark SP; Iwata, So; Fishwick, Colin WG; Johnson, A Peter; Cameron, Alexander D; Henderson, Peter JF

    2014-01-01

    The hydantoin transporter Mhp1 is a sodium-coupled secondary active transport protein of the nucleobase-cation-symport family and a member of the widespread 5-helix inverted repeat superfamily of transporters. The structure of Mhp1 was previously solved in three different conformations providing insight into the molecular basis of the alternating access mechanism. Here, we elucidate detailed events of substrate binding, through a combination of crystallography, molecular dynamics, site-directed mutagenesis, biochemical/biophysical assays, and the design and synthesis of novel ligands. We show precisely where 5-substituted hydantoin substrates bind in an extended configuration at the interface of the bundle and hash domains. They are recognised through hydrogen bonds to the hydantoin moiety and the complementarity of the 5-substituent for a hydrophobic pocket in the protein. Furthermore, we describe a novel structure of an intermediate state of the protein with the external thin gate locked open by an inhibitor, 5-(2-naphthylmethyl)-L-hydantoin, which becomes a substrate when leucine 363 is changed to an alanine. We deduce the molecular events that underlie acquisition and transport of a ligand by Mhp1. PMID:24952894

  10. Molecular recognition of RhlB and RNase D in the Caulobacter crescentus RNA degradosome

    PubMed Central

    Voss, Jarrod E.; Luisi, Ben F.; Hardwick, Steven W.

    2014-01-01

    The endoribonuclease RNase E is a key enzyme in RNA metabolism for many bacterial species. In Escherichia coli, RNase E contributes to the majority of RNA turnover and processing events, and the enzyme has been extensively characterized as the central component of the RNA degradosome assembly. A similar RNA degradosome assembly has been described in the α-proteobacterium Caulobacter crescentus, with the interacting partners of RNase E identified as the Kreb's cycle enzyme aconitase, a DEAD-box RNA helicase RhlB and the exoribonuclease polynucleotide phosphorylase. Here we report that an additional degradosome component is the essential exoribonuclease RNase D, and its recognition site within RNase E is identified. We show that, unlike its E. coli counterpart, C. crescentus RhlB interacts directly with a segment of the N-terminal catalytic domain of RNase E. The crystal structure of a portion of C. crescentus RNase E encompassing the helicase-binding region is reported. This structure reveals that an inserted segment in the S1 domain adopts an α-helical conformation, despite being predicted to be natively unstructured. We discuss the implications of these findings for the organization and mechanisms of the RNA degradosome. PMID:25389270

  11. Scaffolding Student Learning in Clinical Practice.

    ERIC Educational Resources Information Center

    Spouse, Jenny

    1998-01-01

    A longitudinal study of nursing students showed that without sponsorship by clinical staff students found it difficult to participate and learn. The strategy of scaffolding, building on Vygotsky's Zone of Proximal Development, enables recognition of learning needs and the relationship between theory and practice. (SK)

  12. Association, intrinsic shape, and molecular recognition: Elucidating DNA biophysics through coarse-grained simulation

    NASA Astrophysics Data System (ADS)

    Freeman, Gordon Samuel

    DNA is of central importance in biology as it is responsible for carrying, copying, and translating the genetic code into the building blocks that comprise life. In order to accomplish these tasks, the DNA molecule must be versatile and robust. Indeed, the underlying molecular interactions that allow DNA to execute these tasks are complex and their origins are only beginning to be understood. While experiments are able to elucidate many key biophysical phenomena, there remain many unanswered questions. Molecular simulation is able to shed light on phenomena at the molecular scale and provide information that is missing from experimental views of DNA behavior. In this dissertation I use state-of-the-art coarse-grained DNA models to address two key problems. In the first, metadynamics calculations are employed to uncover the free energy surface of two complimentary DNA strands. This free energy surface takes on the appearance of a hybridization funnel and reveals candidates for intermediate states in the hybridization of short DNA oligomers. Such short oligomers are important building blocks for DNA-driven self-assembly and the mechanism of hybridization in this regime is not well understood. The second problem is that of nucleosome formation. Nucleosomes are the fundamental subunit of genome compaction in the nucleus of a cell. As such, nucleosomes are a key epigenetic factor and affect gene expression and the ability of DNA-binding proteins to locate and bind to the appropriate position in the genome. However, the factors that drive nucleosome positioning are not well understood. While DNA sequence is known to affect nucleosome formation, the mechanism by which it does so has not been established and a number of hypotheses explaining this sequence-dependence exist in the literature. I demonstrate that DNA shape dominates this process with contributions arising from both intrinsic DNA curvature as well as DNA-protein interactions driven by sequence

  13. Molecular Basis of mRNA Cap Recognition by Influenza B Polymerase PB2 Subunit.

    PubMed

    Xie, Lili; Wartchow, Charles; Shia, Steven; Uehara, Kyoko; Steffek, Micah; Warne, Robert; Sutton, James; Muiru, Gladys T; Leonard, Vincent H J; Bussiere, Dirksen E; Ma, Xiaolei

    2016-01-01

    Influenza virus polymerase catalyzes the transcription of viral mRNAs by a process known as "cap-snatching," where the 5'-cap of cellular pre-mRNA is recognized by the PB2 subunit and cleaved 10-13 nucleotides downstream of the cap by the endonuclease PA subunit. Although this mechanism is common to both influenza A (FluA) and influenza B (FluB) viruses, FluB PB2 recognizes a wider range of cap structures including m(7)GpppGm-, m(7)GpppG-, and GpppG-RNA, whereas FluA PB2 utilizes methylated G-capped RNA specifically. Biophysical studies with isolated PB2 cap-binding domain (PB2(cap)) confirm that FluB PB2 has expanded mRNA cap recognition capability, although the affinities toward m(7)GTP are significantly reduced when compared with FluA PB2. The x-ray co-structures of the FluB PB2(cap) with bound cap analogs m(7)GTP and GTP reveal an inverted GTP binding mode that is distinct from the cognate m(7)GTP binding mode shared between FluA and FluB PB2. These results delineate the commonalities and differences in the cap-binding site between FluA and FluB PB2 and will aid structure-guided drug design efforts to identify dual inhibitors of both FluA and FluB PB2. PMID:26559973

  14. The molecular basis for recognition of bacterial ligands at equine TLR2, TLR1 and TLR6.

    PubMed

    Irvine, Katherine Lucy; Hopkins, Lee Jason; Gangloff, Monique; Bryant, Clare Elizabeth

    2013-01-01

    TLR2 recognises bacterial lipopeptides and lipoteichoic acid, and forms heterodimers with TLR1 or TLR6. TLR2 is relatively well characterised in mice and humans, with published crystal structures of human TLR2/1/Pam3CSK4 and murine TLR2/6/Pam2CSK4. Equine TLR4 is activated by a different panel of ligands to human and murine TLR4, but less is known about species differences at TLR2. We therefore cloned equine TLR2, TLR1 and TLR6, which showed over 80% sequence identity with these receptors from other mammals, and performed a structure-function analysis. TLR2/1 and TLR2/6 from both horses and humans dose-dependently responded to lipoteichoic acid from Staphylococcus aureus, with no significant species difference in EC50 at either receptor pair. The EC50 of Pam2CSK4 was the same for equine and human TLR2/6, indicating amino acid differences between the two species' TLRs do not significantly affect ligand recognition. Species differences were seen between the responses to Pam2CSK4 and Pam3CSK4 at TLR2/1. Human TLR2/1, as expected, responded to Pam3CSK4 with greater potency and efficacy than Pam2CSK4. At equine TLR2/1, however, Pam3CSK4 was less potent than Pam2CSK4, with both ligands having similar efficacies. Molecular modelling indicates that the majority of non-conserved ligand-interacting residues are at the periphery of the TLR2 binding pocket and in the ligand peptide-interacting regions, which may cause subtle effects on ligand positioning. These results suggest that there are potentially important species differences in recognition of lipopeptides by TLR2/1, which may affect how the horse deals with bacterial infections. PMID:23826682

  15. The molecular basis for recognition of bacterial ligands at equine TLR2, TLR1 and TLR6

    PubMed Central

    2013-01-01

    TLR2 recognises bacterial lipopeptides and lipoteichoic acid, and forms heterodimers with TLR1 or TLR6. TLR2 is relatively well characterised in mice and humans, with published crystal structures of human TLR2/1/Pam3CSK4 and murine TLR2/6/Pam2CSK4. Equine TLR4 is activated by a different panel of ligands to human and murine TLR4, but less is known about species differences at TLR2. We therefore cloned equine TLR2, TLR1 and TLR6, which showed over 80% sequence identity with these receptors from other mammals, and performed a structure-function analysis. TLR2/1 and TLR2/6 from both horses and humans dose-dependently responded to lipoteichoic acid from Staphylococcus aureus, with no significant species difference in EC50 at either receptor pair. The EC50 of Pam2CSK4 was the same for equine and human TLR2/6, indicating amino acid differences between the two species’ TLRs do not significantly affect ligand recognition. Species differences were seen between the responses to Pam2CSK4 and Pam3CSK4 at TLR2/1. Human TLR2/1, as expected, responded to Pam3CSK4 with greater potency and efficacy than Pam2CSK4. At equine TLR2/1, however, Pam3CSK4 was less potent than Pam2CSK4, with both ligands having similar efficacies. Molecular modelling indicates that the majority of non-conserved ligand-interacting residues are at the periphery of the TLR2 binding pocket and in the ligand peptide-interacting regions, which may cause subtle effects on ligand positioning. These results suggest that there are potentially important species differences in recognition of lipopeptides by TLR2/1, which may affect how the horse deals with bacterial infections. PMID:23826682

  16. Molecular Recognition of Fluorine Impacts Substrate Selectivity in the Fluoroacetyl-CoA Thioesterase FlK

    PubMed Central

    2015-01-01

    The fluoroacetate-producing bacterium Streptomyces cattleya has evolved a fluoroacetyl-CoA thioesterase (FlK) that exhibits a remarkably high level of discrimination for its cognate substrate compared to the cellularly abundant analogue acetyl-CoA, which differs only by the absence of the fluorine substitution. A major determinant of FlK specificity derives from its ability to take advantage of the unique properties of fluorine to enhance the reaction rate, allowing fluorine discrimination under physiological conditions where both substrates are likely to be present at saturating concentrations. Using a combination of pH–rate profiles, pre-steady-state kinetic experiments, and Taft analysis of wild-type and mutant FlKs with a set of substrate analogues, we explore the role of fluorine in controlling the enzyme acylation and deacylation steps. Further analysis of chiral (R)- and (S)-[2H1]fluoroacetyl-CoA substrates demonstrates that a kinetic isotope effect (1.7 ± 0.2) is observed for only the (R)-2H1 isomer, indicating that deacylation requires recognition of the prochiral fluoromethyl group to position the α-carbon for proton abstraction. Taken together, the selectivity for the fluoroacetyl-CoA substrate appears to rely not only on the enhanced polarization provided by the electronegative fluorine substitution but also on molecular recognition of fluorine in both formation and breakdown of the acyl-enzyme intermediate to control active site reactivity. These studies provide insights into the basis of fluorine selectivity in a naturally occurring enzyme–substrate pair, with implications for drug design and the development of fluorine-selective biocatalysts. PMID:24635371

  17. Nanosilica-based molecularly imprinted polymer nanoshell for specific recognition and determination of rhodamine B in red wine and beverages.

    PubMed

    Long, Zerong; Xu, Weiwei; Lu, Yi; Qiu, Hongdeng

    2016-09-01

    A new and facile rhodamine B (RhB)-imprinted polymer nanoshell coating for SiO2 nanoparticles was readily prepared by a combination of silica gel modification and molecular surface imprinting. The RhB-imprinted polymers (RhB-MIPs) were characterized by Fourier transform infrared spectroscopy, scanning electron microscopy, and UV-vis spectroscopy; the binding properties and selectivity of these MIPs were investigated in detail. The uniformly imprinted nanoparticles displayed a rather thin shell thickness (23nm) with highly effective recognition sites, showing homogenous distribution and monolayer adsorption. The maximum MIP adsorption capacity (Qm) was as high as 45.2mgg(-1), with an adsorption equilibrium time of about 15min at ambient temperature. Dynamic rebinding experiments showed that chemical adsorption is crucial for RhB binding to RhB-MIPs. The adsorption isotherm for RhB-MIPs binding could also be described by the Langmuir equation at different temperatures and pH values. Increasing temperature led to an enhanced Qm, a decreased dissociation constant (K'd), and a more negative free energy (ΔG), indicating that adsorption is favored at higher temperatures. Moreover, the adsorption capacity of RhB was remarkably affected by pH. At pH>7, the adsorption of RhB was driven by hydrogen bonding interactions, while at pH<7 electrostatic forces were dominant. Additionally, the MIPs also showed specific recognition of RhB from the standard mixture solution containing five structurally analogs. This method was also successfully employed to determine RhB content in red wine and beverages using three levels of spiking, with recoveries in the range of 91.6-93.1% and relative standard deviations lower than 4.1%. PMID:27372912

  18. Multiscaled exploration of coupled folding and binding of an intrinsically disordered molecular recognition element in measles virus nucleoprotein.

    PubMed

    Wang, Yong; Chu, Xiakun; Longhi, Sonia; Roche, Philippe; Han, Wei; Wang, Erkang; Wang, Jin

    2013-10-01

    Numerous relatively short regions within intrinsically disordered proteins (IDPs) serve as molecular recognition elements (MoREs). They fold into ordered structures upon binding to their partner molecules. Currently, there is still a lack of in-depth understanding of how coupled binding and folding occurs in MoREs. Here, we quantified the unbound ensembles of the α-MoRE within the intrinsically disordered C-terminal domain of the measles virus nucleoprotein. We developed a multiscaled approach by combining a physics-based and an atomic hybrid model to decipher the mechanism by which the α-MoRE interacts with the X domain of the measles virus phosphoprotein. Our multiscaled approach led to remarkable qualitative and quantitative agreements between the theoretical predictions and experimental results (e.g., chemical shifts). We found that the free α-MoRE rapidly interconverts between multiple discrete partially helical conformations and the unfolded state, in accordance with the experimental observations. We quantified the underlying global folding-binding landscape. This leads to a synergistic mechanism in which the recognition event proceeds via (minor) conformational selection, followed by (major) induced folding. We also provided evidence that the α-MoRE is a compact molten globule-like IDP and behaves as a downhill folder in the induced folding process. We further provided a theoretical explanation for the inherent connections between "downhill folding," "molten globule," and "intrinsic disorder" in IDP-related systems. Particularly, we proposed that binding and unbinding of IDPs proceed in a stepwise way through a "kinetic divide-and-conquer" strategy that confers them high specificity without high affinity. PMID:24043820

  19. Molecular cloning of matrix Gla protein: implications for substrate recognition by the vitamin K-dependent gamma-carboxylase.

    PubMed Central

    Price, P A; Fraser, J D; Metz-Virca, G

    1987-01-01

    Matrix Gla protein (MGP), a low molecular weight protein found in bone, dentin, and cartilage, contains 5 residues of the vitamin K-dependent amino acid gamma-carboxyglutamic acid (Gla). We have used antibodies raised against MGP and oligonucleotide probes to screen a lambda gt11 cDNA library constructed from the rat osteosarcoma cells (line ROS 17/2) that had been pretreated with 1 alpha,25-dihydroxyvitamin D3. By sequencing several cloned cDNAs, we established a 523-base-pair sequence that predicts an 84-residue mature MGP and a 19-residue hydrophobic signal peptide. The 84-residue mature rat MGP predicted from the cDNA sequence has an additional 5 residues at its C terminus (-Arg-Arg-Gly-Ala-Lys) not seen in the sequence of MGP isolated from bovine bone. The structure of rat MGP provides insight into the mechanisms by which the vitamin K-dependent gamma-carboxylase recognizes substrate. The present studies show that MGP, unlike other vitamin K-dependent proteins, lacks a propeptide. The absence of an MGP propeptide demonstrates that gamma-carboxylation and secretion of vitamin K-dependent proteins need not be linked to the presence of a propeptide or to its proteolytic removal. The propeptides of other vitamin K-dependent proteins are structurally homologous, and there is evidence that this homologous propeptide domain is important to substrate recognition by the gamma-carboxylase. Mature MGP has a sequence segment (residues 15-30) that is homologous to the propeptide of other vitamin K-dependent proteins and probably serves the same role in gamma-carboxylase recognition. Rat MGP also has a second sequence that has recently been identified in all known vitamin K-dependent vertebrate proteins, the invariant unit Glu-Xaa-Xaa-Xaa-Glu-Xaa-Cys (EXXXEXC). Since the glutamic residues in this unit are sites of gamma-carboxylation, it has been suggested that the EXXXEXC unit could allow the gamma-carboxylase to discriminate between substrate and product. The

  20. Molecular recognition of ketamine by a subset of olfactory G protein–coupled receptors

    PubMed Central

    Saven, Jeffery G.; Matsunami, Hiroaki; Eckenhoff, Roderic G.

    2015-01-01

    Ketamine elicits various neuropharmacological effects, including sedation, analgesia, general anesthesia, and antidepressant activity. Through an in vitro screen, we identified four mouse olfactory receptors (ORs) that responded to ketamine. In addition to their presence in the olfactory epithelium, these G protein (heterotrimeric guanine nucleotide–binding protein)–coupled receptors (GPCRs) are distributed throughout the central nervous system. To better understand the molecular basis of the interactions between ketamine and ORs, we used sequence comparison and molecular modeling to design mutations that (i) increased, reduced, or abolished ketamine responsiveness in responding receptors, and (ii) rendered non-responding receptors responsive to ketamine. We showed that olfactory sensory neurons (OSNs) that expressed distinct ORs responded to ketamine in vivo, suggesting that ORs may serve as functional targets for ketamine. The ability to both abolish and introduce responsiveness to ketamine in GPCRs enabled us to identify and confirm distinct interaction loci in the binding site, which suggested a signature ketamine-binding pocket that may guide exploration of additional receptors for this general anesthetic drug. PMID:25829447

  1. Preparation of magnetic molecularly imprinted polymer for selective recognition of resveratrol in wine.

    PubMed

    Chen, Fang-Fang; Xie, Xiao-Yu; Shi, Yan-Ping

    2013-07-26

    The magnetic molecularly imprinted polymers (MMIPs) for resveratrol were prepared by using surface molecular imprinting technique with a super paramagnetic core-shell nanoparticle as a supporter. Rhapontigenin, which is the analogues of resveratrol, was selected as dummy template molecules to avoid the leakage of trace amount of resveratrol. Acrylamide and ethylene glycol dimethacrylate were chosen as functional monomers and cross-linker, respectively. The obtained MMIPs were characterized by using scanning electron microscopy, Fourier transform infrared spectrum, X-ray diffraction and vibrating sample magnetometer. High performance liquid chromatography was used to analyze the target analytes. The resulting MMIPs exhibited high saturation magnetization of 53.14emug(-1) leading to the fast separation. The adsorption test showed that the MMIPs had high adsorption capacity for resveratrol and contained homogeneous binding sites. The MMIPs were employed as adsorbent of solid phase extraction for determination of resveratrol in real wine samples, and the recoveries of spiked samples ranged from 79.3% to 90.6% with the limit of detection of 4.42ngmL(-1). The prepared MMIPs could be employed to selectively pre-concentrate and determine resveratrol from wine samples. PMID:23481473

  2. p53 dynamics upon response element recognition explored by molecular simulations

    PubMed Central

    Terakawa, Tsuyoshi; Takada, Shoji

    2015-01-01

    p53 is a representative transcription factor that activates multiple target genes. To realize stimulus-dependent specificities, p53 has to recognize targets with structural variety, of which molecular mechanisms are largely unknown. Here, we conducted a series of long-time scale (totally more than 100-ms) coarse-grained molecular dynamics simulations, uncovering structure and dynamics of full-length p53 tetramer that recognizes its response element (RE). We obtained structures of a full-length p53 tetramer that binds to the RE, which is strikingly different from the structure of p53 at search. These structures are not only consistent with previous low-resolution or partial structural information, but also give access to previously unreachable detail, such as the preferential distribution of intrinsically disordered regions, the contacts between core domains, the DNA bending, and the connectivity of linker regions. We also explored how the RE variation affects the structure of the p53-RE complex. Further analysis of simulation trajectories revealed how p53 finds out the RE and how post-translational modifications affect the search mechanism. PMID:26596470

  3. Phage display selection of tight specific binding variants from a hyperthermostable Sso7d scaffold protein library.

    PubMed

    Zhao, Ning; Schmitt, Margaret A; Fisk, John D

    2016-04-01

    Antibodies, the quintessential biological recognition molecules, are not ideal for many applications because of their large size, complex modifications, and thermal and chemical instability. Identifying alternative scaffolds that may be evolved into tight, specific binding molecules with improved physical properties is of increasing interest, particularly for biomedical applications in resource-limited environments. Hyperthermophilic organisms, such as Sulfolobus solfataricus, are an attractive source of highly stable proteins that may serve as starting points for alternative molecular recognition scaffolds. We describe the first application of phage display to identify binding proteins based on the S. solfataricus protein Sso7d scaffold. Sso7d is a small cysteine-free DNA-binding protein (approximately 7 kDa, 63 amino acids), with a melting temperature of nearly 100 °C. Tight-binding Sso7d variants were selected for a diverse set of protein targets from a 10(10) member library, demonstrating the versatility of the scaffold. These Sso7d variants are able to discriminate among closely related human, bovine and rabbit serum albumins. Equilibrium dissociation constants in the nanomolar to low micromolar range were measured via competitive ELISA. Importantly, the Sso7d variants continue to bind their targets in the absence of the phage context. Furthermore, phage-displayed Sso7d variants retain their binding affinity after exposure to temperatures up to 70 °C. Taken together, our results suggest that the Sso7d scaffold will be a complementary addition to the range of non-antibody scaffold proteins that may be utilized in phage display. Variants of hyperthermostable binding proteins have potential applications in diagnostics and therapeutics for environments with extreme conditions of storage and deployment. PMID:26835881

  4. Molecular recognition using receptor-free nanomechanical infrared spectroscopy based on a quantum cascade laser

    PubMed Central

    Kim, Seonghwan; Lee, Dongkyu; Liu, Xunchen; Van Neste, Charles; Jeon, Sangmin; Thundat, Thomas

    2013-01-01

    Speciation of complex mixtures of trace explosives presents a formidable challenge for sensors that rely on chemoselective interfaces due to the unspecific nature of weak intermolecular interactions. Nanomechanical infrared (IR) spectroscopy provides higher selectivity in molecular detection without using chemoselective interfaces by measuring the photothermal effect of adsorbed molecules on a thermally sensitive microcantilever. In addition, unlike conventional IR spectroscopy, the detection sensitivity is drastically enhanced by increasing the IR laser power, since the photothermal signal comes from the absorption of IR photons and nonradiative decay processes. By using a broadly tunable quantum cascade laser for the resonant excitation of molecules, we increased the detection sensitivity by one order of magnitude compared to the use of a conventional IR monochromator. Here, we demonstrate the successful speciation and quantification of picogram levels of ternary mixtures of similar explosives (trinitrotoluene (TNT), cyclotrimethylene trinitramine (RDX), and pentaerythritol tetranitrate (PETN)) using nanomechanical IR spectroscopy. PMID:23346368

  5. Recognition of dengue virus protein using epitope-mediated molecularly imprinted film.

    PubMed

    Tai, Dar-Fu; Lin, Chung-Yin; Wu, Tzong-Zeng; Chen, Li-Kuang

    2005-08-15

    Molecularly imprinted film was fabricated in the presence of a pentadecapeptide onto a quartz crystal microbalance (QCM) chip. This 15-mer peptide has been known as the linear epitope of the dengue virus NS1 protein. Imprinting resulted in an increased polymer affinity toward the corresponding templates but also to the virus protein. Direct detection of the dengue virus protein was achieved quantitatively. The QCM chip response to the NS1 protein was obtained using epitope-mediated imprinting demonstrating a comparable frequency shift in chips immobilized with monoclonal antibodies. The binding effect was further enhanced and confirmed using a monoclonal antibody to form a sandwich with the MIP-NS1 protein complex on the chip. No pretreatment was required. PMID:16097751

  6. Molecular recognition using receptor-free nanomechanical infrared spectroscopy based on a quantum cascade laser.

    PubMed

    Kim, Seonghwan; Lee, Dongkyu; Liu, Xunchen; Van Neste, Charles; Jeon, Sangmin; Thundat, Thomas

    2013-01-01

    Speciation of complex mixtures of trace explosives presents a formidable challenge for sensors that rely on chemoselective interfaces due to the unspecific nature of weak intermolecular interactions. Nanomechanical infrared (IR) spectroscopy provides higher selectivity in molecular detection without using chemoselective interfaces by measuring the photothermal effect of adsorbed molecules on a thermally sensitive microcantilever. In addition, unlike conventional IR spectroscopy, the detection sensitivity is drastically enhanced by increasing the IR laser power, since the photothermal signal comes from the absorption of IR photons and nonradiative decay processes. By using a broadly tunable quantum cascade laser for the resonant excitation of molecules, we increased the detection sensitivity by one order of magnitude compared to the use of a conventional IR monochromator. Here, we demonstrate the successful speciation and quantification of picogram levels of ternary mixtures of similar explosives (trinitrotoluene (TNT), cyclotrimethylene trinitramine (RDX), and pentaerythritol tetranitrate (PETN)) using nanomechanical IR spectroscopy. PMID:23346368

  7. One-Step Facile Surface Engineering of Hydrophobic Nanocrystals with Designer Molecular Recognition

    PubMed Central

    Chen, Tao; Öçsoy, Ismail; Yuan, Quan; Wang, Ruowen; You, Mingxu; Zhao, Zilong; Song, Erqun; Zhang, Xiaobing; Tan, Weihong

    2013-01-01

    High quality nanocrystals have demonstrated substantial potential for biomedical applications. However, being generally hydrophobic, their use has been greatly limited by complicated and inefficient surface engineering that often fails to yield biocompatible nanocrystals with minimal aggregation in biological fluids and active targeting toward specific biomolecules. Using chimeric DNA molecules, we developed a one-step facile surface engineering method for hydrophobic Nanocrystals. The procedure is simple and versatile, generating individual nanocrystals with multiple ligands. In addition, the resulting nanocrystals can actively and specifically target various molecular addresses, varying from nucleic acids to cancer cells. Together, the strategy developed here holds great promise in generating critical technologies needed for biomedical applications of nanocrystals. PMID:22793667

  8. Aptamer-functionalized superporous hydrogels for sequestration and release of growth factors regulated via molecular recognition.

    PubMed

    Battig, Mark R; Huang, Yike; Chen, Niancao; Wang, Yong

    2014-09-01

    While the discovery of highly potent biologics has led to the development of promising therapies for various human diseases, biologics can cause severe toxicity if delivered inappropriately. Thus, great efforts have been made to synthesize polymeric systems for safe and efficient delivery of biologics. However, the application of polymeric delivery systems is often limited by problems such as harsh reaction conditions, low drug sequestration efficiency, and difficult drug release regulation. This study was aimed at developing a superporous material system with a hydrogel and an aptamer to overcome these challenges. The results have shown that the superporous hydrogel is capable of instantaneously and fully sequestering a large amount of growth factors, owing to the presence of superporous architectures and aptamers. Moreover, the sequestering and loading procedure does not involve any harsh conditions. The release kinetics of growth factors can be molecularly modulated by either changing the binding affinity of the aptamer or by using a triggering effector. Therefore, this study presents a promising superporous material for the delivery of highly potent biologics such as growth factors for clinical applications. PMID:24954732

  9. Molecular Recognition of Human Liver Cancer Cells Using DNA Aptamers Generated via Cell-SELEX

    PubMed Central

    Zhang, Liqin; Delgado, Stefanie; Champanhac, Carole; Cansiz, Sena; Wu, Cuichen; Shan, Hong; Tan, Weihong

    2015-01-01

    Most clinical cases of liver cancer cannot be diagnosed until they have evolved to an advanced stage, thus resulting in high mortality. It is well recognized that the implementation of early detection methods and the development of targeted therapies for liver cancer are essential to reducing the high mortality rates associated with this disease. To achieve these goals, molecular probes capable of recognizing liver cancer cell-specific targets are needed. Here we describe a panel of aptamers able to distinguish hepatocarcinoma from normal liver cells. The aptamers, which were selected by cell-based SELEX (Systematic Evolution of Ligands by Exponential Enrichment), have Kd values in the range of 64-349 nM toward the target human hepatoma cell HepG2, and also recognize ovarian cancer cells and lung adenocarcinoma. The proteinase treatment experiment indicated that all aptamers could recognize target HepG2 cells through surface proteins. This outcome suggested that these aptamers could be used as potential probes for further research in cancer studies, such as developing early detection assays, targeted therapies, and imaging agents, as well as for the investigation of common membrane proteins in these distinguishable cancers. PMID:25938802

  10. Molecularly imprinted polymer doped with Hectorite for selective recognition of sinomenine hydrochloride.

    PubMed

    Zhang, W; Fu, H L; Li, X Y; Zhang, H; Wang, N; Li, W; Zhang, X X

    2016-01-01

    In this work, a new and facile method was introduced to prepare molecularly imprinted polymers (MIPs) based on nano clay hectorite (Hec) for sinomenine hydrochloride (SM) analysis. Hec was firstly dissolved in distilled water in order to swell adequately, followed by a common precipitation polymerization with SM as the template, methacrylic acid as monomer, ethylene glycol dimethacrylate as a crosslinker and 2,2-azobisisobutyronitrile as an initiator. Hec@SM-MIPs were characterized by Fourier transform infrared spectrometer, transmission electron microscopy, scanning electron microscopy, energy dispersive X-ray spectroscopy and X-ray diffraction. The maximum binding capacity of Hec@SM-MIPs, SM-MIPs and non-imprinted polymers (NIPs) (Hec@NIPs) was 57.4, 16.8 and 11.6 mg/g, respectively. The reason for this result may be that Hec@SM-MIPs have more binding sites and imprinted cavities for template molecule. Equilibrium data were described by the Langmuir and Freundlich isotherm models. The results showed that the Hec@SM-MIPs adsorption data correlated better with the Langmuir equation than the Freundlich equation under the studied concentration range. In vitro drug release experiment, Hec@SM-MIPs have a better ability to control SM release than SM-MIPs. Therefore, Hec@SM-MIPs were successfully applied to extraction of SM and used as the materials for drug delivery system. PMID:26614052

  11. Stoichiometric molecularly imprinted polymers for the recognition of anti-cancer pro-drug tegafur.

    PubMed

    Mattos Dos Santos, Paula; Hall, Andrew J; Manesiotis, Panagiotis

    2016-05-15

    Molecularly imprinted polymers (MIPs) targeting tegafur, an anti-cancer 5-fluorouracil pro-drug, have been prepared by stoichiometric imprinting using 2,6-bis(acrylamido)pyridine (BAAPy) as the functional monomer. Solution association between tegafur and BAAPy was studied by (1)H NMR titration, which confirmed the formation of 1:1 complexes with an affinity constant of 574±15M(-1) in CDCl3. Evaluation of the synthesised materials by HPLC and equilibrium rebinding experiments revealed high selectivity of the imprinted polymer for the pro-drug vs. 5-fluorouracil and other competing analytes, with maximum imprinting factors of 25.3 and a binding capacity of 45.1μmolg(-1). The synthesised imprinted polymer was employed in solid-phase extraction of the pro-drug using an optimised protocol that included a simple wash with the porogen used in the preparation of the material. Tegafur recoveries of up to 96% were achieved from aqueous samples and 92% from urine samples spiked with the template and three competing analytes. The results demonstrate the potential of the prepared polymers in the pre-concentration of tegafur from biological samples, which could be an invaluable tool in the monitoring of patient compliance and drug uptake and excretion. PMID:26711233

  12. Hypovirus molecular biology: from Koch's postulates to host self-recognition genes that restrict virus transmission.

    PubMed

    Dawe, Angus L; Nuss, Donald L

    2013-01-01

    The idea that viruses can be used to control fungal diseases has been a driving force in mycovirus research since the earliest days. Viruses in the family Hypoviridae associated with reduced virulence (hypovirulence) of the chestnut blight fungus, Cryphonectria parasitica, have held a prominent place in this research. This has been due in part to the severity of the chestnut blight epidemics in North America and Europe and early reports of hypovirulence-mediated mitigation of disease in European forests and successful application for control of chestnut blight in chestnut orchards. A more recent contributing factor has been the development of a hypovirus/C. parasitica experimental system that has overcome many of the challenges associated with mycovirus research, stemming primarily from the exclusive intracellular lifestyle shared by all mycoviruses. This chapter will focus on hypovirus molecular biology with an emphasis on the development of the hypovirus/C. parasitica experimental system and its contributions to fundamental and practical advances in mycovirology and the broader understanding of virus-host interactions and fungal pathogenesis. PMID:23498905

  13. Molecular Basis of Phosphatidylinositol 4-Phosphate and ARF1 GTPase Recognition by the FAPP1 Pleckstrin Homology (PH) Domain

    SciTech Connect

    He, J.; Heroux, A.; Scott, J. L.; Roy, S.; Lenoir, M.; Overduin, M.; Stahelin, R. V.; Kutateladze, T. G.

    2011-05-27

    Four-phosphate-adaptor protein 1 (FAPP1) regulates secretory transport from the trans-Golgi network (TGN) to the plasma membrane. FAPP1 is recruited to the Golgi through binding of its pleckstrin homology (PH) domain to phosphatidylinositol 4-phosphate (PtdIns(4)P) and a small GTPase ADP-ribosylation factor 1 (ARF1). Despite the critical role of FAPP1 in membrane trafficking, the molecular basis of its dual function remains unclear. Here, we report a 1.9 {angstrom} resolution crystal structure of the FAPP1 PH domain and detail the molecular mechanisms of the PtdIns(4)P and ARF1 recognition. The FAPP1 PH domain folds into a seven-stranded {beta}-barrel capped by an {alpha}-helix at one edge, whereas the opposite edge is flanked by three loops and the {beta}4 and {beta}7 strands that form a lipid-binding pocket within the {beta}-barrel. The ARF1-binding site is located on the outer side of the {beta}-barrel as determined by NMR resonance perturbation analysis, mutagenesis, and measurements of binding affinities. The two binding sites have little overlap, allowing FAPP1 PH to associate with both ligands simultaneously and independently. Binding to PtdIns(4)P is enhanced in an acidic environment and is required for membrane penetration and tubulation activity of FAPP1, whereas the GTP-bound conformation of the GTPase is necessary for the interaction with ARF1. Together, these findings provide structural and biochemical insight into the multivalent membrane anchoring by the PH domain that may augment affinity and selectivity of FAPP1 toward the TGN membranes enriched in both PtdIns(4)P and GTP-bound ARF1.

  14. Molecular Basis of Phosphatidylinositol 4-Phosphate and ARF1 GTPase Recognition by the FAPP1 Pleckstrin Homology (PH) Domain*

    PubMed Central

    He, Ju; Scott, Jordan L.; Heroux, Annie; Roy, Siddhartha; Lenoir, Marc; Overduin, Michael; Stahelin, Robert V.; Kutateladze, Tatiana G.

    2011-01-01

    Four-phosphate-adaptor protein 1 (FAPP1) regulates secretory transport from the trans-Golgi network (TGN) to the plasma membrane. FAPP1 is recruited to the Golgi through binding of its pleckstrin homology (PH) domain to phosphatidylinositol 4-phosphate (PtdIns(4)P) and a small GTPase ADP-ribosylation factor 1 (ARF1). Despite the critical role of FAPP1 in membrane trafficking, the molecular basis of its dual function remains unclear. Here, we report a 1.9 Å resolution crystal structure of the FAPP1 PH domain and detail the molecular mechanisms of the PtdIns(4)P and ARF1 recognition. The FAPP1 PH domain folds into a seven-stranded β-barrel capped by an α-helix at one edge, whereas the opposite edge is flanked by three loops and the β4 and β7 strands that form a lipid-binding pocket within the β-barrel. The ARF1-binding site is located on the outer side of the β-barrel as determined by NMR resonance perturbation analysis, mutagenesis, and measurements of binding affinities. The two binding sites have little overlap, allowing FAPP1 PH to associate with both ligands simultaneously and independently. Binding to PtdIns(4)P is enhanced in an acidic environment and is required for membrane penetration and tubulation activity of FAPP1, whereas the GTP-bound conformation of the GTPase is necessary for the interaction with ARF1. Together, these findings provide structural and biochemical insight into the multivalent membrane anchoring by the PH domain that may augment affinity and selectivity of FAPP1 toward the TGN membranes enriched in both PtdIns(4)P and GTP-bound ARF1. PMID:21454700

  15. Molecular basis of phosphatidylinositol 4-phosphate and ARF1 GTPase recognition by the FAPP1 pleckstrin homology (PH) domain.

    PubMed

    He, Ju; Scott, Jordan L; Heroux, Annie; Roy, Siddhartha; Lenoir, Marc; Overduin, Michael; Stahelin, Robert V; Kutateladze, Tatiana G

    2011-05-27

    Four-phosphate-adaptor protein 1 (FAPP1) regulates secretory transport from the trans-Golgi network (TGN) to the plasma membrane. FAPP1 is recruited to the Golgi through binding of its pleckstrin homology (PH) domain to phosphatidylinositol 4-phosphate (PtdIns(4)P) and a small GTPase ADP-ribosylation factor 1 (ARF1). Despite the critical role of FAPP1 in membrane trafficking, the molecular basis of its dual function remains unclear. Here, we report a 1.9 Å resolution crystal structure of the FAPP1 PH domain and detail the molecular mechanisms of the PtdIns(4)P and ARF1 recognition. The FAPP1 PH domain folds into a seven-stranded β-barrel capped by an α-helix at one edge, whereas the opposite edge is flanked by three loops and the β4 and β7 strands that form a lipid-binding pocket within the β-barrel. The ARF1-binding site is located on the outer side of the β-barrel as determined by NMR resonance perturbation analysis, mutagenesis, and measurements of binding affinities. The two binding sites have little overlap, allowing FAPP1 PH to associate with both ligands simultaneously and independently. Binding to PtdIns(4)P is enhanced in an acidic environment and is required for membrane penetration and tubulation activity of FAPP1, whereas the GTP-bound conformation of the GTPase is necessary for the interaction with ARF1. Together, these findings provide structural and biochemical insight into the multivalent membrane anchoring by the PH domain that may augment affinity and selectivity of FAPP1 toward the TGN membranes enriched in both PtdIns(4)P and GTP-bound ARF1. PMID:21454700

  16. Interfacial molecular interactions based on the conformation recognition between the insoluble antitumor drug AD-1 and DSPC.

    PubMed

    Yin, Tian; Cao, Xiuxiu; Liu, Xiaolin; Wang, Jian; Shi, Caihong; Su, Jia; Zhang, Yu; Gou, Jingxin; He, Haibing; Guo, Haiyan; Tang, Xing; Zhao, Yuqing

    2016-10-01

    In this study, molecular interactions between the anti-cancer agent 20(R)-25-methoxyl-dammarane-3β, 12β, 20-triol (AD-1) and phospholipid 1,2-Distearoyl-sn-glycero-3-phosphocholine (DSPC) were investigated using the Langmuir film balance technique. The characteristics of binary Langmuir monolayers consisting of DSPC and AD-1 were conducted on the basis of the surface pressure-area per molecule (π-A) isotherms. It was found that the drug was able to become efficiently inserted into preformed DSPC monolayers, indicating a preferential interaction between AD-1 and DSPC. For the examined lateral pressure at 20mN/m, the largest negative values of ΔGex were found for the AD-1/DSPC monolayer, which should be the most stable. Based on the calculated values of ΔGex, we found that the AD-1/DSPC systems exhibited the best mixed characteristics when the molar fraction of the AD-1 was 0.8; at that relative concentration, the AD-1 molecules can mix better and interact with the phospholipid molecules. In addition, the drug-DSPC binary supramolecular structure was also deposited on the mica plates as shown by atomic force microscopy (AFM). Finally, molecular docking calculations explained satisfactorily that, based on the conformations interactions (conformation recognition), even at an AD-1/DSPC molar ratio as high as 8:2, the interfacial stabilization of the AD-1/DSPC system was fairly strong due to hydrophobic interactions. A higher loading capacity of DSPC might be possible, as it is associated with a more flexible geometrical environment, which allows these supramolecular structures to accept larger increases in drug loading upon steric binding. PMID:27469574

  17. Molecular recognition of naphthalene diimide ligands by telomeric quadruplex-DNA: the importance of the protonation state and mediated hydrogen bonds.

    PubMed

    Spinello, A; Barone, G; Grunenberg, J

    2016-01-28

    In depth Monte Carlo conformational scans in combination with molecular dynamics (MD) simulations and electronic structure calculations were applied in order to study the molecular recognition process between tetrasubstituted naphthalene diimide (ND) guests and G-quadruplex (G4) DNA receptors. ND guests are a promising class of telomere stabilizers due to which they are used in novel anticancer therapeutics. Though several ND guests have been studied experimentally in the past, the protonation state under physiological conditions is still unclear. Based on chemical intuition, in the case of N-methyl-piperazine substitution, different protonation states are possible and might play a crucial role in the molecular recognition process by G4-DNA. Depending on the proton concentration, different nitrogen atoms of the N-methyl-piperazine might (or might not) be protonated. This fact was considered in our simulation in terms of a case by case analysis, since the process of molecular recognition is determined by possible donor or acceptor positions. The results of our simulations show that the electrostatic interactions between the ND ligands and the G4 receptor are maximized in the case of the protonation of the terminal nitrogen atoms, forming compact ND G4 complexes inside the grooves. The influence of different protonation states in terms of the ability to form hydrogen bonds with the sugar-phosphate backbone, as well as the importance of mediated vs. direct hydrogen bonding, was analyzed in detail by MD and relaxed force constant (compliance constant) simulations. PMID:26733046

  18. Molecular recognition of the antiretroviral drug abacavir: towards the development of a novel carbazole-based fluorosensor.

    PubMed

    Idzik, Krzysztof Ryszard; Cywinski, Piotr J; Cranfield, Charles G; Mohr, Gerhard J; Beckert, Rainer

    2011-05-01

    Due to their optical and electro-conductive attributes, carbazole derivatives are interesting materials for a large range of biosensor applications. In this study, we present the synthesis routes and fluorescence evaluation of newly designed carbazole fluorosensors that, by modification with uracil, have a special affinity for antiretroviral drugs via either Watson-Crick or Hoogsteen base pairing. To an N-octylcarbazole-uracil compound, four different groups were attached, namely thiophene, furane, ethylenedioxythiophene, and another uracil; yielding four different derivatives. Photophysical properties of these newly obtained derivatives are described, as are their interactions with the reverse transcriptase inhibitors such as abacavir, zidovudine, lamivudine and didanosine. The influence of each analyte on biosensor fluorescence was assessed on the basis of the Stern-Volmer equation and represented by Stern-Volmer constants. Consequently we have demonstrated that these structures based on carbazole, with a uracil group, may be successfully incorporated into alternative carbazole derivatives to form biosensors for the molecular recognition of antiretroviral drugs. PMID:21222147

  19. Sialyllactose in Viral Membrane Gangliosides Is a Novel Molecular Recognition Pattern for Mature Dendritic Cell Capture of HIV-1

    PubMed Central

    Contreras, F.-Xabier; Rodriguez-Plata, Maria T.; Glass, Bärbel; Erkizia, Itziar; Prado, Julia G.; Casas, Josefina; Fabriàs, Gemma; Kräusslich, Hans-Georg; Martinez-Picado, Javier

    2012-01-01

    HIV-1 is internalized into mature dendritic cells (mDCs) via an as yet undefined mechanism with subsequent transfer of stored, infectious virus to CD4+ T lymphocytes. Thus, HIV-1 subverts a DC antigen capture mechanism to promote viral spread. Here, we show that gangliosides in the HIV-1 membrane are the key molecules for mDC uptake. HIV-1 virus-like particles and liposomes mimicking the HIV-1 lipid composition were shown to use a common internalization pathway and the same trafficking route within mDCs. Hence, these results demonstrate that gangliosides can act as viral attachment factors, in addition to their well known function as cellular receptors for certain viruses. Furthermore, the sialyllactose molecule present in specific gangliosides was identified as the determinant moiety for mDC HIV-1 uptake. Thus, sialyllactose represents a novel molecular recognition pattern for mDC capture, and may be crucial both for antigen presentation leading to immunity against pathogens and for succumbing to subversion by HIV-1. PMID:22545022

  20. Mitotic regulator Mis18β interacts with and specifies the centromeric assembly of molecular chaperone holliday junction recognition protein (HJURP).

    PubMed

    Wang, Jianyu; Liu, Xing; Dou, Zhen; Chen, Liang; Jiang, Hao; Fu, Chuanhai; Fu, Guosheng; Liu, Dan; Zhang, Jiancun; Zhu, Tongge; Fang, Jingwen; Zang, Jianye; Cheng, Jinke; Teng, Maikun; Ding, Xia; Yao, Xuebiao

    2014-03-21

    The centromere is essential for precise and equal segregation of the parental genome into two daughter cells during mitosis. CENP-A is a unique histone H3 variant conserved in eukaryotic centromeres. The assembly of CENP-A to the centromere is mediated by Holliday junction recognition protein (HJURP) in early G1 phase. However, it remains elusive how HJURP governs CENP-A incorporation into the centromere. Here we show that human HJURP directly binds to Mis18β, a component of the Mis18 complex conserved in the eukaryotic kingdom. A minimal region of HJURP for Mis18β binding was mapped to residues 437-460. Depletion of Mis18β by RNA interference dramatically impaired HJURP recruitment to the centromere, indicating the importance of Mis18β in HJURP loading. Interestingly, phosphorylation of HJURP by CDK1 weakens its interaction with Mis18β, consistent with the notion that assembly of CENP-A to the centromere is achieved after mitosis. Taken together, these data define a novel molecular mechanism underlying the temporal regulation of CENP-A incorporation into the centromere by accurate Mis18β-HJURP interaction. PMID:24519934

  1. Mitotic Regulator Mis18β Interacts with and Specifies the Centromeric Assembly of Molecular Chaperone Holliday Junction Recognition Protein (HJURP)*

    PubMed Central

    Wang, Jianyu; Liu, Xing; Dou, Zhen; Chen, Liang; Jiang, Hao; Fu, Chuanhai; Fu, Guosheng; Liu, Dan; Zhang, Jiancun; Zhu, Tongge; Fang, Jingwen; Zang, Jianye; Cheng, Jinke; Teng, Maikun; Ding, Xia; Yao, Xuebiao

    2014-01-01

    The centromere is essential for precise and equal segregation of the parental genome into two daughter cells during mitosis. CENP-A is a unique histone H3 variant conserved in eukaryotic centromeres. The assembly of CENP-A to the centromere is mediated by Holliday junction recognition protein (HJURP) in early G1 phase. However, it remains elusive how HJURP governs CENP-A incorporation into the centromere. Here we show that human HJURP directly binds to Mis18β, a component of the Mis18 complex conserved in the eukaryotic kingdom. A minimal region of HJURP for Mis18β binding was mapped to residues 437–460. Depletion of Mis18β by RNA interference dramatically impaired HJURP recruitment to the centromere, indicating the importance of Mis18β in HJURP loading. Interestingly, phosphorylation of HJURP by CDK1 weakens its interaction with Mis18β, consistent with the notion that assembly of CENP-A to the centromere is achieved after mitosis. Taken together, these data define a novel molecular mechanism underlying the temporal regulation of CENP-A incorporation into the centromere by accurate Mis18β-HJURP interaction. PMID:24519934

  2. dsRNA-protein interactions studied by molecular dynamics techniques. Unravelling dsRNA recognition by DCL1.

    PubMed

    Drusin, Salvador I; Suarez, Irina P; Gauto, Diego F; Rasia, Rodolfo M; Moreno, Diego M

    2016-04-15

    Double stranded RNA (dsRNA) participates in several biological processes, where RNA molecules acquire secondary structure inside the cell through base complementarity. The double stranded RNA binding domain (dsRBD) is one of the main protein folds that is able to recognize and bind to dsRNA regions. The N-terminal dsRBD of DCL1 in Arabidopsis thaliana (DCL1-1), in contrast to other studied dsRBDs, lacks a stable structure, behaving as an intrinsically disordered protein. DCL1-1 does however recognize dsRNA by acquiring a canonical fold in the presence of its substrate. Here we present a detailed modeling and molecular dynamics study of dsRNA recognition by DCL1-1. We found that DCL1-1 forms stable complexes with different RNAs and we characterized the residues involved in binding. Although the domain shows a binding loop substantially shorter than other homologs, it can still interact with the dsRNA and results in bending of the dsRNA A-type helix. Furthermore, we found that R8, a non-conserved residue located in the first dsRNA binding region, recognizes preferentially mismatched base pairs. We discuss our findings in the context of the function of DCL1-1 within the microRNA processing complex. PMID:26987516

  3. Molecular basis for H3K36me3 recognition by the Tudor domain of PHF1

    PubMed Central

    Musselman, Catherine A.; Avvakumov, Nikita; Watanabe, Reiko; Abraham, Christopher G.; Lalonde, Marie-Eve; Hong, Zehui; Allen, Christopher; Roy, Siddhartha; Nuñez, James K.; Nickoloff, Jac; Kulesza, Caroline A.; Yasui, Akira; Côté, Jacques; Kutateladze, Tatiana G.

    2013-01-01

    The PHD finger protein 1 (PHF1) is essential in epigenetic regulation and genome maintenance. Here, we demonstrate that the Tudor domain of human PHF1 binds to histone H3 trimethylated at Lys36 (H3K36me3). We report a 1.9 Å resolution crystal structure of the Tudor domain in complex with H3K36me3 and describe the molecular mechanism of H3K36me3 recognition using NMR analysis. Binding of PHF1 to H3K36me3 inhibits the ability of the Polycomb PRC2 complex to methylate H3K27 in vitro and in vivo. Laser micro-irradiation data reveal that PHF1 is transiently recruited to DNA double-strand breaks (DSBs), and PHF1 mutants impaired in the H3K36me3 interaction exhibit reduced retention at DSB sites. Together, our findings suggest that PHF1 can mediate deposition of the repressive H3K27me3 mark and acts as an early DNA damage response cofactor. PMID:23142980

  4. Recombinant protein scaffolds for tissue engineering.

    PubMed

    Werkmeister, Jerome A; Ramshaw, John A M

    2012-02-01

    New biological materials for tissue engineering are now being developed using common genetic engineering capabilities to clone and express a variety of genetic elements that allow cost-effective purification and scaffold fabrication from these recombinant proteins, peptides or from chimeric combinations of these. The field is limitless as long as the gene sequences are known. The utility is dependent on the ease, product yield and adaptability of these protein products to the biomedical field. The development of recombinant proteins as scaffolds, while still an emerging technology with respect to commercial products, is scientifically superior to current use of natural materials or synthetic polymer scaffolds, in terms of designing specific structures with desired degrees of biological complexities and motifs. In the field of tissue engineering, next generation scaffolds will be the key to directing appropriate tissue regeneration. The initial period of biodegradable synthetic scaffolds that provided shape and mechanical integrity, but no biological information, is phasing out. The era of protein scaffolds offers distinct advantages, particularly with the combination of powerful tools of molecular biology. These include, for example, the production of human proteins of uniform quality that are free of infectious agents and the ability to make suitable quantities of proteins that are found in low quantity or are hard to isolate from tissue. For the particular needs of tissue engineering scaffolds, fibrous proteins like collagens, elastin, silks and combinations of these offer further advantages of natural well-defined structural scaffolds as well as endless possibilities of controlling functionality by genetic manipulation. PMID:22262725

  5. The molecular bases of δ/αβ T cell–mediated antigen recognition

    PubMed Central

    Pellicci, Daniel G.; Uldrich, Adam P.; Le Nours, Jérôme; Ross, Fiona; Chabrol, Eric; Eckle, Sidonia B.G.; de Boer, Renate; Lim, Ricky T.; McPherson, Kirsty; Besra, Gurdyal; Howell, Amy R.; Moretta, Lorenzo; McCluskey, James; Heemskerk, Mirjam H.M.; Gras, Stephanie

    2014-01-01

    αβ and γδ T cells are disparate T cell lineages that can respond to distinct antigens (Ags) via the use of the αβ and γδ T cell Ag receptors (TCRs), respectively. Here we characterize a population of human T cells, which we term δ/αβ T cells, expressing TCRs comprised of a TCR-δ variable gene (Vδ1) fused to joining α and constant α domains, paired with an array of TCR-β chains. We demonstrate that these cells, which represent ∼50% of all Vδ1+ human T cells, can recognize peptide- and lipid-based Ags presented by human leukocyte antigen (HLA) and CD1d, respectively. Similar to type I natural killer T (NKT) cells, CD1d-lipid Ag-reactive δ/αβ T cells recognized α-galactosylceramide (α-GalCer); however, their fine specificity for other lipid Ags presented by CD1d, such as α-glucosylceramide, was distinct from type I NKT cells. Thus, δ/αβTCRs contribute new patterns of Ag specificity to the human immune system. Furthermore, we provide the molecular bases of how δ/αβTCRs bind to their targets, with the Vδ1-encoded region providing a major contribution to δ/αβTCR binding. Our findings highlight how components from αβ and γδTCR gene loci can recombine to confer Ag specificity, thus expanding our understanding of T cell biology and TCR diversity. PMID:25452463

  6. Molecular Mechanism of Flocculation Self-Recognition in Yeast and Its Role in Mating and Survival

    PubMed Central

    Goossens, Katty V. Y.; Ielasi, Francesco S.; Nookaew, Intawat; Stals, Ingeborg; Alonso-Sarduy, Livan; Daenen, Luk; Van Mulders, Sebastiaan E.; Stassen, Catherine; van Eijsden, Rudy G. E.; Siewers, Verena; Delvaux, Freddy R.; Kasas, Sandor; Nielsen, Jens; Devreese, Bart

    2015-01-01

    ABSTRACT We studied the flocculation mechanism at the molecular level by determining the atomic structures of N-Flo1p and N-Lg-Flo1p in complex with their ligands. We show that they have similar ligand binding mechanisms but distinct carbohydrate specificities and affinities, which are determined by the compactness of the binding site. We characterized the glycans of Flo1p and their role in this binding process and demonstrate that glycan-glycan interactions significantly contribute to the cell-cell adhesion mechanism. Therefore, the extended flocculation mechanism is based on the self-interaction of Flo proteins and this interaction is established in two stages, involving both glycan-glycan and protein-glycan interactions. The crucial role of calcium in both types of interaction was demonstrated: Ca2+ takes part in the binding of the carbohydrate to the protein, and the glycans aggregate only in the presence of Ca2+. These results unify the generally accepted lectin hypothesis with the historically first-proposed “Ca2+-bridge” hypothesis. Additionally, a new role of cell flocculation is demonstrated; i.e., flocculation is linked to cell conjugation and mating, and survival chances consequently increase significantly by spore formation and by introduction of genetic variability. The role of Flo1p in mating was demonstrated by showing that mating efficiency is increased when cells flocculate and by differential transcriptome analysis of flocculating versus nonflocculating cells in a low-shear environment (microgravity). The results show that a multicellular clump (floc) provides a uniquely organized multicellular ultrastructure that provides a suitable microenvironment to induce and perform cell conjugation and mating. PMID:25873380

  7. Molecular Dynamics Analysis of Antibody Recognition and Escape by Human H1N1 Influenza Hemagglutinin

    PubMed Central

    Ieong, Pek; Amaro, Rommie E.; Li, Wilfred W.

    2015-01-01

    The antibody immunoglobulin (Ig) 2D1 is effective against the 1918 hemagglutinin (HA) and also known to cross-neutralize the 2009 pandemic H1N1 influenza HA through a similar epitope. However, the detailed mechanism of neutralization remains unclear. We conducted molecular dynamics (MD) simulations to study the interactions between Ig-2D1 and the HAs from the 1918 pandemic flu (A/South Carolina/1/1918, 18HA), the 2009 pandemic flu (A/California/04/2009, 09HA), a 2009 pandemic flu mutant (A/California/04/2009, 09HA_mut), and the 2006 seasonal flu (A/Solomon Islands/3/2006, 06HA). MM-PBSA analyses suggest the approximate free energy of binding (ΔG) between Ig-2D1 and 18HA is −74.4 kcal/mol. In comparison with 18HA, 09HA and 06HA bind Ig-2D1 ∼6 kcal/mol (ΔΔG) weaker, and the 09HA_mut bind Ig-2D1 only half as strong. We also analyzed the contributions of individual epitope residues using the free-energy decomposition method. Two important salt bridges are found between the HAs and Ig-2D1. In 09HA, a serine-to-asparagine mutation coincided with a salt bridge destabilization, hydrogen bond losses, and a water pocket formation between 09HA and Ig-2D1. In 09HA_mut, a lysine-to-glutamic-acid mutation leads to the loss of both salt bridges and destabilizes interactions with Ig-2D1. Even though 06HA has a similar ΔG to 09HA, it is not recognized by Ig-2D1 in vivo. Because 06HA contains two potential glycosylation sites that could mask the epitope, our results suggest that Ig-2D1 may be active against 06HA only in the absence of glycosylation. Overall, our simulation results are in good agreement with observations from biological experiments and offer novel mechanistic insights, to our knowledge, into the immune escape of the influenza virus. PMID:26039171

  8. Contig-Layout-Authenticator (CLA): A Combinatorial Approach to Ordering and Scaffolding of Bacterial Contigs for Comparative Genomics and Molecular Epidemiology

    PubMed Central

    Shaik, Sabiha; Kumar, Narender; Lankapalli, Aditya K.; Tiwari, Sumeet K.; Baddam, Ramani; Ahmed, Niyaz

    2016-01-01

    A wide variety of genome sequencing platforms have emerged in the recent past. High-throughput platforms like Illumina and 454 are essentially adaptations of the shotgun approach generating millions of fragmented single or paired sequencing reads. To reconstruct whole genomes, the reads have to be assembled into contigs, which often require further downstream processing. The contigs can be directly ordered according to a reference, scaffolded based on paired read information, or assembled using a combination of the two approaches. While the reference-based approach appears to mask strain-specific information, scaffolding based on paired-end information suffers when repetitive elements longer than the size of the sequencing reads are present in the genome. Sequencing technologies that produce long reads can solve the problems associated with repetitive elements but are not necessarily easily available to researchers. The most common high-throughput technology currently used is the Illumina short read platform. To improve upon the shortcomings associated with the construction of draft genomes with Illumina paired-end sequencing, we developed Contig-Layout-Authenticator (CLA). The CLA pipeline can scaffold reference-sorted contigs based on paired reads, resulting in better assembled genomes. Moreover, CLA also hints at probable misassemblies and contaminations, for the users to cross-check before constructing the consensus draft. The CLA pipeline was designed and trained extensively on various bacterial genome datasets for the ordering and scaffolding of large repetitive contigs. The tool has been validated and compared favorably with other widely-used scaffolding and ordering tools using both simulated and real sequence datasets. CLA is a user friendly tool that requires a single command line input to generate ordered scaffolds. PMID:27248146

  9. Scaffolder - software for manual genome scaffolding

    PubMed Central

    2012-01-01

    Background The assembly of next-generation short-read sequencing data can result in a fragmented non-contiguous set of genomic sequences. Therefore a common step in a genome project is to join neighbouring sequence regions together and fill gaps. This scaffolding step is non-trivial and requires manually editing large blocks of nucleotide sequence. Joining these sequences together also hides the source of each region in the final genome sequence. Taken together these considerations may make reproducing or editing an existing genome scaffold difficult. Methods The software outlined here, “Scaffolder,” is implemented in the Ruby programming language and can be installed via the RubyGems software management system. Genome scaffolds are defined using YAML - a data format which is both human and machine-readable. Command line binaries and extensive documentation are available. Results This software allows a genome build to be defined in terms of the constituent sequences using a relatively simple syntax. This syntax further allows unknown regions to be specified and additional sequence to be used to fill known gaps in the scaffold. Defining the genome construction in a file makes the scaffolding process reproducible and easier to edit compared with large FASTA nucleotide sequences. Conclusions Scaffolder is easy-to-use genome scaffolding software which promotes reproducibility and continuous development in a genome project. Scaffolder can be found at http://next.gs. PMID:22640820

  10. Electrospinning and crosslinking of low-molecular-weight poly(trimethylene carbonate-co-(L)-lactide) as an elastomeric scaffold for vascular engineering.

    PubMed

    Dargaville, Bronwin L; Vaquette, Cédryck; Rasoul, Firas; Cooper-White, Justin J; Campbell, Julie H; Whittaker, Andrew K

    2013-06-01

    The growth of suitable tissue to replace natural blood vessels requires a degradable scaffold material that is processable into porous structures with appropriate mechanical and cell growth properties. This study investigates the fabrication of degradable, crosslinkable prepolymers of l-lactide-co-trimethylene carbonate into porous scaffolds by electrospinning. After crosslinking by γ-radiation, dimensionally stable scaffolds were obtained with up to 56% trimethylene carbonate incorporation. The fibrous mats showed Young's moduli closely matching human arteries (0.4-0.8MPa). Repeated cyclic extension yielded negligible change in mechanical properties, demonstrating the potential for use under dynamic physiological conditions. The scaffolds remained elastic and resilient at 30% strain after 84days of degradation in phosphate buffer, while the modulus and ultimate stress and strain progressively decreased. The electrospun mats are mechanically superior to solid films of the same materials. In vitro, human mesenchymal stem cells adhered to and readily proliferated on the three-dimensional fiber network, demonstrating that these polymers may find use in growing artificial blood vessels in vivo. PMID:23416575

  11. Designing and preparation of novel alkaloid-imprinted membrane with grafting type and its molecular recognition characteristic and permselectivity.

    PubMed

    Gao, Baojiao; Zhang, Liqin; Li, Yanbin

    2016-09-01

    A novel polysulfone-based molecularly imprinted membrane (MIM) with graft type (designated as GMIM) was successfully prepared by a combination of film-forming method of immersion-precipitation phase transformation with molecule surface-imprinting technique. The porous asymmetry membrane of chloromethylated polysulfone (CMPSF) was first prepared by a phase inversion method, and then the CMPSF membrane was amination-modified with ethanediamine as reagent, resulting aminated polysulfone membrane AMPSF, on whose surface primary amino groups were contained. Then the graft-polymerization of methacrylic acid (MAA) was realized by initiating of the surface-initiating system of -NH2/S2O8(-), obtaining the grafted membrane PSF-g-PMAA. After PSF-g-PMAA membrane adsorbed matrine, the crosslinking reaction of the grafted PMAA was allowed to be carried out with ethylene glycol diglycidyl ether (EGDE) as crosslinker, resulting in the matrine imprinted membrane with graft type, GMIM. The binding characteristics of the imprinted membrane GMIM, the permeability and separation property for matrine were investigated in depth. The experimental results show that the imprinted membrane consists of a thin imprinted layer, a thin skin layer containing channels at nanoscale and a support layer with macroporous structure. The imprinted membrane GMIM has specific recognition selectivity and excellent binding affinity for matrine, and its selectivity coefficient for matrine relative to cytisine is 4.85. More importantly, the imprinted membrane can produce good "gate effect" because of its own structure characteristic, so that it has fine permselectivity for the template, matrine molecule. The separation coefficient of the imprinted membrane GMIM for matrine relative to cytisine as a contrast reaches up to 5.9, displaying the excellent performance of a selectively permeable membrane. PMID:27207062

  12. Radial scan of the molecular electrostatic potential of RNA double helices: an application to the enzyme-tRNA recognition.

    PubMed

    Marín, Ray M; Agudelo, William A; Daza C, Edgar E

    2008-10-01

    We introduced a method to characterize quantitatively the molecular electrostatic potential (MEP) of the minor and major grooves of base pairs located at nucleic acid double helices. By means of a radial MEP scan, we obtained a n-tuple of potential values corresponding to each groove, which can be analyzed by plotting the MEP values as a function of the angle in the radial scan. We studied base pairs of two different tRNAs, relevant in the recognition process with their cognate aminoacyl tRNA synthetases (aaRSs), in order to correlate their electrostatic behavior with the corresponding aminoacylation activity. We analyzed the first three base pairs of the Escherichia coli tRNA(Ala) acceptor stem, finding several cases where the MEP profiles obtained from the plots are in agreement with the reported aminoacylation activities. Additionally, a non-hierarchical clustering performed over the MEP n-tuples resulted in meaningful classifications that correlate with the activity and with the predicted stereochemistry of the reaction. We also studied the first two base pairs of the E. coli tRNA(Thr) acceptor stem but constraining the analysis to the angle intervals that seem relevant for the binding sites of the enzyme. These intervals were deduced from the ThrRS-tRNA(Thr) complex crystal structure. In this case, we also found a good agreement between the MEP profiles and the activity, supporting the idea that the tRNA identity elements function is to allow an optimal electrostatic complementarity between the aminoacyl-tRNA synthetase and the tRNA. PMID:18586541

  13. Molecular bases for the recognition of short peptide substrates and cysteine-directed modifications of human insulin-degrading enzyme

    PubMed Central

    Malito, Enrico; Ralat, Luis A.; Manolopoulou, Marika; Tsay, Julie L.; Wadlington, Natasha L.; Tang, Wei-Jen

    2009-01-01

    Insulin degrading enzyme (IDE) utilizes a large catalytic chamber to selectively bind and degrade peptide substrates such as insulin and amyloid β (Aβ). Tight interactions with substrates occur at an exosite located ~30Å away from the catalytic center that anchors the N-terminus of substrates to facilitate binding and subsequent cleavages at the catalytic site. However, IDE also degrades peptide substrates that are too short to occupy both the catalytic site and the exosite simultaneously. Here, we use kinins as a model system to address the kinetics and regulation of human IDE with short peptides. IDE specifically degrades bradykinin and kallidin at the Pro/Phe site. A 1.9Å crystal structure of bradykinin-bound IDE reveals the binding of bradykinin to the exosite, and not to the catalytic site. In agreement with observed high Km values, this suggests low affinity of bradykinin for IDE. This structure also provides the molecular basis on how the binding of short peptides at the exosite could regulate substrate recognition. We also found that human IDE is potently inhibited by physiologically relevant concentrations of S-nitrosylation and oxidation agents. Cysteine-directed modifications play a key role, since an IDE mutant devoid of all thirteen cysteines is insensitive to the inhibition by S-nitroso-glutathione, hydrogen peroxide, or N-ethylmaleimide. Specifically, cysteine 819 of human IDE is located inside the catalytic chamber pointing towards an extended hydrophobic pocket and is critical for the inactivation. Thiol-directed modification of this residue likely causes local structural perturbation to reduce substrate binding and catalysis. PMID:18986166

  14. Surface Characterization of Extracellular Matrix Scaffolds

    PubMed Central

    Brown, Bryan N.; Barnes, Christopher A.; Kasick, Rena T.; Michel, Roger; Gilbert, Thomas W.; Beer-Stolz, Donna; Castner, David G.; Ratner, Buddy D.; Badylak, Stephen F.

    2009-01-01

    Extracellular matrix (ECM) scaffolds prepared from different tissue sources or using different methods have been demonstrated to have distinctive effects upon cell adhesion patterns and the ability to support and maintain differentiated phenotypes. It is unknown whether the molecular composition or the ultrastructure of the ECM plays a greater role in determining the phenotype of the cells with which it comes into contact. However, when implanted, the topology and ligand landscape of the material will determine the host molecules that bind and the type and behavior of cells that mediate the host response. Therefore, a comprehensive understanding of surface characteristics is essential in the design of scaffolds for specific clinical applications. The surface characteristics of ECM scaffolds derived from porcine urinary bladder, small intestine, and liver as well as the effects of two commonly used methods of chemical cross-linking upon UBM were investigated. Electron microscopy and time of flight secondary ion mass spectroscopy were used to examine the surface characteristics of the scaffolds. The results show that ECM scaffolds have unique morphologic and structural properties which are dependant on the organ or tissue from which the scaffold is harvested. Furthermore, the results show that the surface characteristics of an ECM scaffold are changed through chemical cross-linking. PMID:19828192

  15. Biomimetic nanoclay scaffolds for bone tissue engineering

    NASA Astrophysics Data System (ADS)

    Ambre, Avinash Harishchandra

    Tissue engineering offers a significant potential alternative to conventional methods for rectifying tissue defects by evoking natural regeneration process via interactions between cells and 3D porous scaffolds. Imparting adequate mechanical properties to biodegradable scaffolds for bone tissue engineering is an important challenge and extends from molecular to macroscale. This work focuses on the use of sodium montmorillonite (Na-MMT) to design polymer composite scaffolds having enhanced mechanical properties along with multiple interdependent properties. Materials design beginning at the molecular level was used in which Na-MMT clay was modified with three different unnatural amino acids and further characterized using Fourier Transform Infrared (FTIR) spectroscopy, X-ray diffraction (XRD). Based on improved bicompatibility with human osteoblasts (bone cells) and intermediate increase in d-spacing of MMT clay (shown by XRD), 5-aminovaleric acid modified clay was further used to prepare biopolymer (chitosan-polygalacturonic acid complex) scaffolds. Osteoblast proliferation in biopolymer scaffolds containing 5-aminovaleric acid modified clay was similar to biopolymer scaffolds containing hydroxyapatite (HAP). A novel process based on biomineralization in bone was designed to prepare 5-aminovaleric acid modified clay capable of imparting multiple properties to the scaffolds. Bone-like apatite was mineralized in modified clay and a novel nanoclay-HAP hybrid (in situ HAPclay) was obtained. FTIR spectroscopy indicated a molecular level organic-inorganic association between the intercalated 5-aminovaleric acid and mineralized HAP. Osteoblasts formed clusters on biopolymer composite films prepared with different weight percent compositions of in situ HAPclay. Human MSCs formed mineralized nodules on composite films and mineralized extracellular matrix (ECM) in composite scaffolds without the use of osteogenic supplements. Polycaprolactone (PCL), a synthetic polymer, was

  16. Bio-inspired artificial functional photocatalyst: biomimetic enzyme-like TiO2/reduced graphene oxide nanocomposite with excellent molecular recognition ability

    NASA Astrophysics Data System (ADS)

    Li, Wentao; Pei, Xule; Deng, Fang; Luo, Xubiao; Li, Fengcong; Xiao, Yong

    2015-05-01

    An enzyme-like TiO2/reduced graphene oxide (enzyme-TiO2/rGO) nanocomposite with molecular recognition ability was fabricated by biomimicking the geometrical and chemical complementation of the enzyme and substrate. The anatase TiO2 nanocrystals were densely dispersed on rGO nanosheets with close interfacial contacts. With geometrical and chemical matching of target molecules and memorized cavities, the adsorption capacity of enzyme-TiO2/rGO nanocomposites for 4-nitrophenol (4.71 mg g-1) is about six times that of control TiO2/rGO without the enzyme-like feature (0.79 mg g-1), and the enzyme-TiO2/rGO shows a relative selectivity coefficient of 7.24. Moreover, enzyme-TiO2/rGO exhibits molecular recognitive photocatalytic degradation for a particular contaminant. The results demonstrate that enzyme-substrate recognition provides a convenient and powerful basis on which to biomimic and construct efficient photocatalysts with high selectivity.

  17. SOFT-MI: a novel microfabrication technique integrating soft-lithography and molecular imprinting for tissue engineering applications.

    PubMed

    Vozzi, Giovanni; Morelli, Ilaria; Vozzi, Federico; Andreoni, Chiara; Salsedo, Elisabetta; Morachioli, Annagiulia; Giusti, Paolo; Ciardelli, Gianluca

    2010-08-01

    An innovative approach has been employed for the realization of bioactive scaffolds able to mimic the in vivo cellular microenvironment for tissue engineering applications. This method is based on the combination of molecular imprinting and soft-lithography technology to enhance cellular adhesion and to guide cell growth and proliferation due to presence of highly specific recognition sites of selected biomolecules on a well-defined polymeric microstructure. In this article polymethylmethacrylate (PMMA) scaffolds have been realized by using poly(dimethylsiloxane) (PDMS) microstructured molds imprinted with FITC-albumin and TRITC-lectin. In addition gelatin, an adhesion protein, was employed for the molecular imprinting of polymeric scaffolds for cellular tests. The most innovative aspect of this research was the molecular imprinting of whole cells for the development of substrates able to enhance the cell adhesion processes. PMID:20564617

  18. Co-assembly of CdTe and Fe3O4 with molecularly imprinted polymer for recognition and separation of endocrine disrupting chemicals

    NASA Astrophysics Data System (ADS)

    Chang, Limin; Chen, Shaona; Chu, Jia; Li, Xin

    2013-11-01

    In this study, we present a general protocol to fabricate imprinting matrix co-loaded with CdTe quantum dots and Fe3O4 nanoparticles for the recognition of endocrine disrupting chemicals (EDCs). The resultant composites were characterized by transmission electron microscopy, fluorescence spectroscopy, and energy dispersive spectroscopy. The materials have been demonstrated to be characterized with spherical shape with a saturation magnetization value of 1.7 emu g-1. Furthermore, the rebinding experiments show that the resultant materials have greater affinity and selectivity towards p-nitrophenol (model EDCs) over structurally related compounds. We believe that the effective method proposed in this work might provide a platform to prepare magnetic and fluorescent molecularly imprinted polymers for the recognition and separation of EDCs.

  19. Molecular recognition in a diverse set of protein-ligand interactions studied with molecular dynamics simulations and end-point free energy calculations.

    PubMed

    Wang, Bo; Li, Liwei; Hurley, Thomas D; Meroueh, Samy O

    2013-10-28

    End-point free energy calculations using MM-GBSA and MM-PBSA provide a detailed understanding of molecular recognition in protein-ligand interactions. The binding free energy can be used to rank-order protein-ligand structures in virtual screening for compound or target identification. Here, we carry out free energy calculations for a diverse set of 11 proteins bound to 14 small molecules using extensive explicit-solvent MD simulations. The structure of these complexes was previously solved by crystallography and their binding studied with isothermal titration calorimetry (ITC) data enabling direct comparison to the MM-GBSA and MM-PBSA calculations. Four MM-GBSA and three MM-PBSA calculations reproduced the ITC free energy within 1 kcal·mol(-1) highlighting the challenges in reproducing the absolute free energy from end-point free energy calculations. MM-GBSA exhibited better rank-ordering with a Spearman ρ of 0.68 compared to 0.40 for MM-PBSA with dielectric constant (ε = 1). An increase in ε resulted in significantly better rank-ordering for MM-PBSA (ρ = 0.91 for ε = 10), but larger ε significantly reduced the contributions of electrostatics, suggesting that the improvement is due to the nonpolar and entropy components, rather than a better representation of the electrostatics. The SVRKB scoring function applied to MD snapshots resulted in excellent rank-ordering (ρ = 0.81). Calculations of the configurational entropy using normal-mode analysis led to free energies that correlated significantly better to the ITC free energy than the MD-based quasi-harmonic approach, but the computed entropies showed no correlation with the ITC entropy. When the adaptation energy is taken into consideration by running separate simulations for complex, apo, and ligand (MM-PBSAADAPT), there is less agreement with the ITC data for the individual free energies, but remarkably good rank-ordering is observed (ρ = 0.89). Interestingly, filtering MD snapshots by prescoring

  20. Heteromeric assembled polypeptidic artificial hydrolases with a six-helical bundle scaffold.

    PubMed

    Bai, Yu; Ling, Yanbo; Shi, Weiguo; Cai, Lifeng; Jia, Qiyan; Jiang, Shibo; Liu, Keliang

    2011-11-25

    Enzyme efficiency results from the cooperation of functional groups in the catalytic site. In order to mimic a natural enzyme, a definite 3D scaffold must be carefully designed so that the functional groups can work cooperatively. During the HIV-1 fusion process, the gp41 N- and C-terminal heptad repeat regions form a coiled-coil six-helical bundle (6HB) that brings the viral and target cell membranes into close proximity for fusion. We used 6HB as the molecular model for a novel scaffold for the design of an artificial enzyme, in which the modified C34 and N36 peptides formed a unique 6HB structure through specific molecular recognition, and the position and orientation of the side-chain groups on this scaffold were predictable. The histidine modified 6HB C34(H13/20)/N36(H15/22) showed enzyme-like hydrolytic activity towards p-nitrophenyl acetate (PNPA; k(cat)/K(M) =3.66 M(-1) s(-1)) through the cooperation of several inter- or intrahelical imidazole groups. Since the catalytic activity of 6HB depends on the C- and N-peptide assembly, either HIV fusion inhibitors that can compete with the formation of catalytic 6HB or denaturants that can destroy the ordered structure were able to modulate its activity. Further engineering of the solvent-exposing face with Glu(-)-Lys(+) salt bridges enhanced the helicity and the stability of 6HB. As a result, the population and stability of cooperative catalytic units increased. In addition, the Glu(-)-Lys(+) -stabilized 6HB SC35(H13/20)/N36(H15/22) had increased catalytic efficiency (k(cat)/K(M) =6.30 M(-1) s(-1)). A unique 6HB system was specifically assembled and provided a scaffold sufficiently stable to mimic the function of enzymes or other biomolecules. PMID:21957084

  1. Molecular recognition of proline tRNA by prolyl-tRNA synthetase from hyperthermophilic archaeon, Aeropyrum pernix K1.

    PubMed

    Yokozawa, Junji; Okamoto, Koji; Kawarabayasi, Yutaka; Kuno, Atsushi; Hasegawa, Tsunemi

    2003-01-01

    To investigate the recognition mechanism of tRNA(Pro) by prolyl-tRNA synthetase from hyperthermophilic archaeon, Aeropyrum pernix K1, various tRNA(Pro) transcripts were prepared by in vitro transcription system. These transcripts were aminoacylated with proline by overexpressed A. pernix prolyl-tRNA synthetase. From prolylation experiments, recognition elements of A. pernix tRNA(Pro) were determined to be G35 and G36 of anticodon, discriminator base A73, and G1-C72 base pair at acceptor stem end. PMID:14510473

  2. [Bone tissue engineering scaffolds].

    PubMed

    Fang, Liru; Weng, Wenjian; Shen, Ge; Han, Gaorong; Santos, J D; Du, Peiyi

    2003-03-01

    Bone tissue engineering may provide an alternative to the repairs to skeletal defects resulting from disease, trauma or surgery. Scaffold has played an important role in bone tissue engineering, which functions as the architecture for bone in growth. In this paper, the authors gave a brief introduction about the requirement of bone tissue engineering scaffold, the key of the design of scaffolds and the current research on this subject. PMID:12744187

  3. Molecular recognition principles and stationary-phase characteristics of topoisomer-selective chemoaffinity materials for chromatographic separation of circular plasmid DNA topoisomers.

    PubMed

    Mahut, Marek; Lindner, Wolfgang; Lämmerhofer, Michael

    2012-01-18

    We recently discovered the molecular recognition capability of a quinine carbamate ligand attached to silica as a powerful chemoaffinity material for the chromatographic separation of circular plasmid topoisomers of different linking numbers. In this paper we develop structure-selectivity relationship studies to figure out the essential structural features for topoisomer recognition. By varying different moieties of the original cinchonan-derived selector, it was shown that intercalation by the quinoline moiety of the ligand as assumed initially as the working hypothesis is not an essential feature for topoisomer recognition during chromatography. We found that the key elements for topoisomer selectivity are the presence of a rigid weak anion-exchange site and a H-donor site separated from each other in a defined distance by a 4-atom spacer. Additionally, incorporation of the weak anion-exchange site into a cyclic ring structure provides greater rigidity of the ligand molecule and turned out to be advantageous, if not mandatory, for (close to) baseline separation. PMID:22191385

  4. A supramolecular hyperbranched polymer based on molecular recognition between benzo-21-crown-7 and secondary ammonium salt.

    PubMed

    Li, Hui; Fan, Xiaodong; Tian, Wei; Zhang, Haitao; Zhang, Wanbin; Yang, Zhen

    2014-12-01

    A novel A2-B3-type supramolecular hyperbranched polymer was prepared based on the benzo-21-crown-7/secondary ammonium salt recognition motif. The resulting supramolecular polymer exhibited a reversible disassembled-assembled process by adding or removing potassium ions. PMID:25315034

  5. Suppression of apoptosis by enhanced protein adsorption on polymer/hydroxyapatite composite scaffolds

    PubMed Central

    Woo, Kyung Mi; Seo, Jihye; Zhang, Ruiyun; Ma, Peter X.

    2007-01-01

    Bone tissue engineering is a promising alternative to bone grafting. Scaffolds play a critical role in tissue engineering. Composite scaffolds made of biodegradable polymers and bone mineral-like inorganic compounds have been reported to be advantageous over plain polymer scaffolds by our group and others. In this study, we compared cellular and molecular events during the early periods of osteoblastic cell culture on poly(l-lactic acid)/hydroxyapatite (PLLA/HAP) composite scaffolds with those on plain PLLA scaffolds, and showed that PLLA/HAP scaffolds improved cell survival over plain PLLA scaffolds. Most cells (MC3T3-E1) on PLLA/HAP scaffolds survived the early culture. In contrast, about 50% of the cells initially adhered to the plain PLLA scaffolds were detached within the first 12 h and showed characteristics of apoptotic cell death, which was confirmed by TUNEL staining and caspase-3 activation. To investigate the mechanisms, we examined the adsorption of serum protein and adhesion molecules to the scaffolds. The PLLA/HAP scaffold adsorbed more than 1.4 times of total serum protein and much greater amounts of serum fibronectin and vitronectin than pure PLLA scaffolds. Similarly, significantly larger amounts of individual adhesion proteins and peptides (fibronectin, vitronectin, RGD, and KRSR) were adsorbed on the PLLA/HAP scaffolds than on the PLLA scaffolds, which resulted in higher cell density on the PLLA/HAP scaffolds. Furthermore, β1 and β3 integrins and phosphorylation of Fak and Akt proteins in the cells on the PLLA/HAP scaffolds were significantly more abundent than those on PLLA scaffolds, which suggest that enhanced adsorption of serum adhesion proteins to PLLA/HAP scaffolds protect the cells from apoptosis possibly through the integrin-Fak-Akt pathway. These results demonstrate that biomimetic composite scaffolds are advantageous for bone tissue engineering. PMID:17320948

  6. A Cyclized Helix-Loop-Helix Peptide as a Molecular Scaffold for the Design of Inhibitors of Intracellular Protein-Protein Interactions by Epitope and Arginine Grafting.

    PubMed

    Fujiwara, Daisuke; Kitada, Hidekazu; Oguri, Masahiro; Nishihara, Toshio; Michigami, Masataka; Shiraishi, Kazunori; Yuba, Eiji; Nakase, Ikuhiko; Im, Haeri; Cho, Sunhee; Joung, Jong Young; Kodama, Seiji; Kono, Kenji; Ham, Sihyun; Fujii, Ikuo

    2016-08-26

    The design of inhibitors of intracellular protein-protein interactions (PPIs) remains a challenge in chemical biology and drug discovery. We propose a cyclized helix-loop-helix (cHLH) peptide as a scaffold for generating cell-permeable PPI inhibitors through bifunctional grafting: epitope grafting to provide binding activity, and arginine grafting to endow cell-permeability. To inhibit p53-HDM2 interactions, the p53 epitope was grafted onto the C-terminal helix and six Arg residues were grafted onto another helix. The designed peptide cHLHp53-R showed high inhibitory activity for this interaction, and computational analysis suggested a binding mode for HDM2. Confocal microscopy of cells treated with fluorescently labeled cHLHp53-R revealed cell membrane penetration and cytosolic localization. The peptide inhibited the growth of HCT116 and LnCap cancer cells. This strategy of bifunctional grafting onto a well-structured peptide scaffold could facilitate the generation of inhibitors for intracellular PPIs. PMID:27467415

  7. New Trends in Inspecting GPCR-ligand Recognition Process: the Contribution of the Molecular Modeling Section (MMS) at the University of Padova.

    PubMed

    Ciancetta, Antonella; Cuzzolin, Alberto; Deganutti, Giuseppe; Sturlese, Mattia; Salmaso, Veronica; Cristiani, Andrea; Sabbadin, Davide; Moro, Stefano

    2016-09-01

    In this review, we present a survey of the recent advances carried out by our research groups in the field of ligand-GPCRs recognition process simulations recently implemented at the Molecular Modeling Section (MMS) of the University of Padova. We briefly describe a platform of tools we have tuned to aid the identification of novel GPCRs binders and the better understanding of their binding mechanisms, based on two extensively used computational techniques such as molecular docking and MD simulations. The developed methodologies encompass: (i) the selection of suitable protocols for docking studies, (ii) the exploration of the dynamical evolution of ligand-protein interaction networks, (iii) the detailed investigation of the role of water molecules upon ligand binding, and (iv) a glance at the way the ligand might go through prior reaching the binding site. PMID:27546048

  8. Hydrogels and scaffolds for immunomodulation

    PubMed Central

    2014-01-01

    For over two decades, immunologists and biomaterials scientists have co-existed in parallel world with the rationale of understanding the molecular profile of immune responses to vaccination, implantation, and treating incurable diseases. Much of the field of biomaterials-based immunotherapy has relied on evaluating model antigens such as chicken egg ovalbumin in mouse models but their relevance to humans has been point of much discussion. Nevertheless, such model antigens have provided important insights about the mechanisms of immune regulation and served as a proof-of-concept for plethora of biomaterials-based vaccines. After years of extensive development of numerous biomaterials for immunomodulation, it is only recently that an experimental scaffold vaccine implanted beneath the skin has begun to use the human model to study the immune responses to cancer vaccination by co-delivering patient-derived tumor lysates and immunomodulatory proteins. If successful, this scaffold vaccine will change the way we approached untreatable cancers, but more importantly, will allow a faster and more rational translation of therapeutic regimes to other cancers, chronic infections, and autoimmune diseases. Most materials reviews have focused on immunomodulatory adjuvants and micro-nano-particles. Here we provide an insight into emerging hydrogel and scaffold based immunomodulatory approaches that continue to demonstrate efficacy against immune associated diseases. PMID:25155610

  9. Hydrogels and scaffolds for immunomodulation.

    PubMed

    Singh, Ankur; Peppas, Nicholas A

    2014-10-01

    For over two decades, immunologists and biomaterials scientists have co-existed in parallel world with the rationale of understanding the molecular profile of immune responses to vaccination, implantation, and treating incurable diseases. Much of the field of biomaterial-based immunotherapy has relied on evaluating model antigens such as chicken egg ovalbumin in mouse models but their relevance to humans has been point of much discussion. Nevertheless, such model antigens have provided important insights into the mechanisms of immune regulation and served as a proof-of-concept for plethora of biomaterial-based vaccines. After years of extensive development of numerous biomaterials for immunomodulation, it is only recently that an experimental scaffold vaccine implanted beneath the skin has begun to use the human model to study the immune responses to cancer vaccination by co-delivering patient-derived tumor lysates and immunomodulatory proteins. If successful, this scaffold vaccine will change the way we approached untreatable cancers, but more importantly, will allow a faster and more rational translation of therapeutic regimes to other cancers, chronic infections, and autoimmune diseases. Most materials reviews have focused on immunomodulatory adjuvants and micro-nano-particles. Here we provide an insight into emerging hydrogel and scaffold based immunomodulatory approaches that continue to demonstrate efficacy against immune associated diseases. PMID:25155610

  10. Recognition and binding of β-lactam antibiotics to bovine serum albumin by frontal affinity chromatography in combination with spectroscopy and molecular docking.

    PubMed

    Li, Qian; Zhang, Tianlong; Bian, Liujiao

    2016-03-01

    Serum albumins are the most abundant carrier proteins in blood plasma and participate in the binding and transportation of various exogenous and endogenous compounds in the body. This work was designed to investigate the recognition and binding of three typical β-lactam antibiotics including penicillin G (Pen G), penicillin V (Pen V) and cefalexin (Cef) with bovine serum albumin (BSA) by frontal affinity chromatography in combination with UV-vis absorption spectra, fluorescence emission spectra, binding site marker competitive experiment and molecular docking under simulated physiological conditions. The results showed that a BSA only bound with one antibiotic molecule in the binding process, and the binding constants for Pen G-BSA, Pen V-BSA and Cef-BSA complexes were 4.22×10(1), 4.86×10(2) and 3.32×10(3) (L/mol), respectively. All the three β-lactam antibiotics were mainly inserted into the subdomain IIA (binding site 1) of BSA by hydrogen bonds and Van der Waals forces. The binding capacity between the antibiotics and BSA was closely related to the functional groups and flexibility of side chains in antibiotics. This study provided an important insight into the molecular recognition and binding interaction of BSA with β-lactam antibiotics, which may be a useful guideline for the innovative clinical medications and new antibiotic designs with effective pharmacological properties. PMID:26882128

  11. Molecular basis for the recognition of cyclic-di-AMP by PstA, a PII-like signal transduction protein.

    PubMed

    Choi, Philip H; Sureka, Kamakshi; Woodward, Joshua J; Tong, Liang

    2015-06-01

    Cyclic-di-AMP (c-di-AMP) is a broadly conserved bacterial second messenger that is of importance in bacterial physiology. The molecular receptors mediating the cellular responses to the c-di-AMP signal are just beginning to be discovered. PstA is a previously uncharacterized PII -like protein which has been identified as a c-di-AMP receptor. PstA is widely distributed and conserved among Gram-positive bacteria in the phylum Firmicutes. Here, we report the biochemical, structural, and functional characterization of PstA from Listeria monocytogenes. We have determined the crystal structures of PstA in the c-di-AMP-bound and apo forms at 1.6 and 2.9 Å resolution, respectively, which provide the molecular basis for its specific recognition of c-di-AMP. PstA forms a homotrimer structure that has overall similarity to the PII protein family which binds ATP. However, PstA is markedly different from PII proteins in the loop regions, and these structural differences mediate the specific recognition of their respective nucleotide ligand. The residues composing the c-di-AMP binding pocket are conserved, suggesting that c-di-AMP recognition by PstA is of functional importance. Disruption of pstA in L. monocytogenes affected c-di-AMP-mediated alterations in bacterial growth and lysis. Overall, we have defined the PstA family as a conserved and specific c-di-AMP receptor in bacteria. PMID:25693966

  12. Molecular basis for the recognition of cyclic-di-AMP by PstA, a PII-like signal transduction protein

    PubMed Central

    Choi, Philip H; Sureka, Kamakshi; Woodward, Joshua J; Tong, Liang

    2015-01-01

    Cyclic-di-AMP (c-di-AMP) is a broadly conserved bacterial second messenger that is of importance in bacterial physiology. The molecular receptors mediating the cellular responses to the c-di-AMP signal are just beginning to be discovered. PstA is a previously uncharacterized PII-like protein which has been identified as a c-di-AMP receptor. PstA is widely distributed and conserved among Gram-positive bacteria in the phylum Firmicutes. Here, we report the biochemical, structural, and functional characterization of PstA from Listeria monocytogenes. We have determined the crystal structures of PstA in the c-di-AMP-bound and apo forms at 1.6 and 2.9 Å resolution, respectively, which provide the molecular basis for its specific recognition of c-di-AMP. PstA forms a homotrimer structure that has overall similarity to the PII protein family which binds ATP. However, PstA is markedly different from PII proteins in the loop regions, and these structural differences mediate the specific recognition of their respective nucleotide ligand. The residues composing the c-di-AMP binding pocket are conserved, suggesting that c-di-AMP recognition by PstA is of functional importance. Disruption of pstA in L. monocytogenes affected c-di-AMP-mediated alterations in bacterial growth and lysis. Overall, we have defined the PstA family as a conserved and specific c-di-AMP receptor in bacteria. PMID:25693966

  13. Molecular Evolution of the CYP2D Subfamily in Primates: Purifying Selection on Substrate Recognition Sites without the Frequent or Long-Tract Gene Conversion

    PubMed Central

    Yasukochi, Yoshiki; Satta, Yoko

    2015-01-01

    The human cytochrome P450 (CYP) 2D6 gene is a member of the CYP2D gene subfamily, along with the CYP2D7P and CYP2D8P pseudogenes. Although the CYP2D6 enzyme has been studied extensively because of its clinical importance, the evolution of the CYP2D subfamily has not yet been fully understood. Therefore, the goal of this study was to reveal the evolutionary process of the human drug metabolic system. Here, we investigate molecular evolution of the CYP2D subfamily in primates by comparing 14 CYP2D sequences from humans to New World monkey genomes. Window analysis and statistical tests revealed that entire genomic sequences of paralogous genes were extensively homogenized by gene conversion during molecular evolution of CYP2D genes in primates. A neighbor-joining tree based on genomic sequences at the nonsubstrate recognition sites showed that CYP2D6 and CYP2D8 genes were clustered together due to gene conversion. In contrast, a phylogenetic tree using amino acid sequences at substrate recognition sites did not cluster the CYP2D6 and CYP2D8 genes, suggesting that the functional constraint on substrate specificity is one of the causes for purifying selection at the substrate recognition sites. Our results suggest that the CYP2D gene subfamily in primates has evolved to maintain the regioselectivity for a substrate hydroxylation activity between individual enzymes, even though extensive gene conversion has occurred across CYP2D coding sequences. PMID:25808902

  14. Hierarchical scaffolding with Bambus.

    PubMed

    Pop, Mihai; Kosack, Daniel S; Salzberg, Steven L

    2004-01-01

    The output of a genome assembler generally comprises a collection of contiguous DNA sequences (contigs) whose relative placement along the genome is not defined. A procedure called scaffolding is commonly used to order and orient these contigs using paired read information. This ordering of contigs is an essential step when finishing and analyzing the data from a whole-genome shotgun project. Most recent assemblers include a scaffolding module; however, users have little control over the scaffolding algorithm or the information produced. We thus developed a general-purpose scaffolder, called Bambus, which affords users significant flexibility in controlling the scaffolding parameters. Bambus was used recently to scaffold the low-coverage draft dog genome data. Most significantly, Bambus enables the use of linking data other than that inferred from mate-pair information. For example, the sequence of a completed genome can be used to guide the scaffolding of a related organism. We present several applications of Bambus: support for finishing, comparative genomics, analysis of the haplotype structure of genomes, and scaffolding of a mammalian genome at low coverage. Bambus is available as an open-source package from our Web site. PMID:14707177

  15. Molecular recognition of tRNA(Pro) by Escherichia coli proline tRNA synthetase in vitro.

    PubMed

    Liu, H; Peterson, R; Kessler, J; Musier-Forsyth, K

    1995-01-11

    In this study, we identify a subset of nucleotides that specify aminoacylation of tRNA(Pro) by Escherichia coli proline tRNA synthetase in vitro. Twenty-two tRNA(Pro) variants were prepared by in vitro transcription and their efficiency of aminoacylation with proline (kcat/KM) was measured. From this analysis, we conclude that recognition elements for tRNA(Pro) aminoacylation by ProRS are located in at least three domains of the tRNA molecule. The largest decreases in the kinetic parameters for aminoacylation resulted from single substitutions at position G72 of the acceptor stem and position G36 of the anticodon. Anticodon nucleotide G35 and position A73 in the acceptor stem were also identified as major recognition elements. Moreover, bases that are believed to be important for maintaining the tertiary structure of the tRNA (G15 and C48) appear to be important for efficient recognition of tRNA(Pro) by ProRS in vitro. PMID:7870582

  16. Molecularly imprinted polymer grafted on polysaccharide microsphere surface by the sol-gel process for protein recognition.

    PubMed

    Li, Feng; Li, Jing; Zhang, Shusheng

    2008-02-15

    An interfacial organic-inorganic hybridization concept was applied to the preparation of a new spherical imprinted material for protein recognition. The functional biopolymer chitosan (CS), shaped as microsphere and high-density cross-linked, constituted of the polysaccharide core for surface imprinting. After the model template protein, bovine serum albumin, was covalently immobilized by forming imine bonds with the functional amine groups of CS, two kinds of organic siloxane (3-aminopropyltrimethoxysiloxane: APTMS, and tetraethoxysiloxane: TEOS) assembled and polymerized on the polysaccharide-protein surface via sol-gel process in aqueous solution at room temperature. After template removal, the protein-imprinted sol-gel surface exhibited a prevalent preference for the template protein in adsorption experiments, as compared with four contrastive proteins. Bioinformatics methods were also employed to investigate the imprinting process and the recognition effect. The influence of siloxane type, pH, siloxane/water ratio on template removal and recognition selectivity was assessed. Under optimized imprinting conditions, a large quantity of well-distributed pores was observed on the immobilized-template imprinted surface. The surface-imprinted adsorbent offered a fast kinetics for template re-adsorption and could be reused. Compared with the imprinted material prepared with free-template, material prepared with immobilized-template possessed higher adsorption capacity towards template protein. Easy preparation of the described imprinted material, high affinity and good reusability make this approach attractive and broadly applicable in biotechnology for down-stream processing and biosensor. PMID:18371777

  17. Cytosine derivatized bis(2,2'-bithienyl)methane molecularly imprinted polymer for selective recognition of 6-thioguanine, an antitumor drug.

    PubMed

    Huynh, Tan-Phat; Wojnarowicz, Agnieszka; Sosnowska, Marta; Srebnik, Simcha; Benincori, Tiziana; Sannicolò, Francesco; D'Souza, Francis; Kutner, Wlodzimierz

    2015-08-15

    A molecularly imprinted polymer (MIP) was designed and synthesized to serve as a functional material for selective recognition of 6-thioguanine (6TG), an antitumor drug. For that, the newly synthesized functional monomer, cytosine-bis(2,2'-bithienyl)-(4-carboxyphenyl)methane ester (Cyt-S4), revealed Watson-Crick type nucleobase pairing of 6TG. Formation of the Cyt-S4 and 6TG complex of the 2:1 stoichiometry was postulated based on the DFT calculations at the B3LYP/3-21G((⁎)) level and experimentally confirmed by fluorescence titration. The molecularly imprinted polymer (MIP) film was deposited by potentiodynamic electropolymerization on a Pt disk electrode as well as on an Au-coated glass slide and on an Au-quartz crystal resonator. The statistical model of formation of this film was successfully simulated by molecular dynamics. Completeness of the subsequent 6TG template extraction from MIP was confirmed by the UV-visible spectroscopy. An imprinting factor of 2.9 for the MIP film was determined by piezoelectric microgravimetry using ECQM. The double-layer capacity and alternating current measurements under flow-injection analysis (FIA) conditions were selected to transduce the 6TG recognition signal into the change of the double-layer capacity dependence on the 6TG concentration in solution for different supporting electrolyte concentrations. Detectability of the resulting chemosensor was 10 µM 6TG for the 0.5 M KF carrier solution in FIA. Selectivity of the chemosensor with respect to common interferences was high, e.g., it exceeded 130 to 2-amino-6-methylmercaptopurine, a 6TG metabolite. PMID:25805629

  18. On the Influence of Crosslinker on Template Complexation in Molecularly Imprinted Polymers: A Computational Study of Prepolymerization Mixture Events with Correlations to Template-Polymer Recognition Behavior and NMR Spectroscopic Studies

    PubMed Central

    Shoravi, Siamak; Olsson, Gustaf D.; Karlsson, Björn C. G.; Nicholls, Ian A.

    2014-01-01

    Aspects of the molecular-level basis for the function of ethylene glycol dimethacrylate and trimethylolproprane trimethacrylate crosslinked methacrylic acid copolymers molecularly imprinted with (S)-propranolol have been studied using a series of all-component and all-atom molecular dynamics studies of the corresponding prepolymerization systems. The crosslinking agents were observed to contribute to template complexation, and the results were contrasted with previously reported template-recognition behavior of the corresponding polymers. Differences in the extent to which the two crosslinkers interacted with the functional monomer were identified, and correlations were made to polymer-ligand recognition behavior and the results of nuclear magnetic resonance spectroscopic studies studies. This study demonstrates the importance of considering the functional monomer–crosslinker interaction when designing molecularly imprinted polymers, and highlights the often neglected general contribution of crosslinker to determining the nature of molecularly imprinted polymer-template selectivity. PMID:24927149

  19. The Interaction between an Acidic Transcriptional Activator and Its Inhibitor: The Molecular Basis of Ga14p Recognition by Ga180p

    SciTech Connect

    Thoden, James B.; Ryan, Louise A.; Reece, Richard J.; Holden, Hazel M.

    2009-02-16

    The GAL genes, which encode the enzymes required for normal galactose metabolism in yeast, are transcriptionally regulated by three proteins: Gal4p, an activator; Gal80p, an inhibitor; and Gal3p, a galactose sensor. These proteins control the switch between inert and active gene expression. The transcriptional activation function of Gal4p is rendered inactive in the presence of Gal80p. Here we present the three-dimensional structure of a complex between the acidic activation domain of Gal4p and Gal80p. The transactivation domain initiates with an extended region of polypeptide chain followed by two turns of an amphipathic alpha-helix. It fits into and across a deep cleft within the Gal80p dimer with the protein-protein interface defined primarily by hydrophobic interactions. A disordered loop in the apo-Gal80p structure (Asp-309 to Ser-316) becomes well-defined upon binding of the transactivation domain. This investigation provides a new molecular scaffold for understanding previous biochemical and genetic studies.

  20. Molecular Basis for the Recognition of Structurally Distinct Autoinducer Mimics by the Pseudomonas aeruginosa LasR Quorum-Sensing Signaling Receptor

    SciTech Connect

    Zou, Yaozhong; Nair, Satish K.

    2010-01-12

    The human pathogen Pseudomonas aeruginosa coordinates the expression of virulence factors using quorum sensing, a signaling cascade triggered by the activation of signal receptors by small-molecule autoinducers. These homoserine lactone autoinducers stabilize their cognate receptors and activate their functions as transcription factors. Because quorum sensing regulates the progression of infection and host immune resistance, significant efforts have been devoted toward the identification of small molecules that disrupt this process. Screening efforts have identified a class of triphenyl compounds that are structurally distinct from the homoserine lactone autoinducer, yet interact specifically and potently with LasR receptor to modulate quorum sensing (Muh et al., 2006a). Here we present the high-resolution crystal structures of the ligand binding domain of LasR in complex with the autoinducer N-3-oxo-dodecanoyl homoserine lactone (1.4 {angstrom} resolution), and with the triphenyl mimics TP-1, TP-3, and TP-4 (to between 1.8 {angstrom} and 2.3 {angstrom} resolution). These crystal structures provide a molecular rationale for understanding how chemically distinct compounds can be accommodated by a highly selective receptor, and provide the framework for the development of novel quorum-sensing regulators, utilizing the triphenyl scaffold.

  1. Proton transfer in organic scaffolds

    NASA Astrophysics Data System (ADS)

    Basak, Dipankar

    This dissertation focuses on the fundamental understanding of the proton transfer process and translating the knowledge into design/development of new organic materials for efficient non-aqueous proton transport. For example, what controls the shuttling of a proton between two basic sites? a) Distance between two groups? or b) the basicity? c) What is the impact of protonation on molecular conformation when the basic sites are attached to rigid scaffolds? For this purpose, we developed several tunable proton sponges and studied proton transfer in these scaffolds theoretically as well as experimentally. Next we moved our attention to understand long-range proton conduction or proton transport. We introduced liquid crystalline (LC) proton conductor based on triphenylene molecule and established that activation energy barrier for proton transport is lower in the LC phase compared to the crystalline phase. Furthermore, we investigated the impact of several critical factors: the choice of the proton transferring groups, mobility of the charge carriers, intrinsic vs. extrinsic charge carrier concentrations and the molecular architectures on long-range proton transport. The outcome of this research will lead to a deeper understanding of non-aqueous proton transfer process and aid the design of next generation proton exchange membrane (PEM) for fuel cell.

  2. Molecular recognition with metal containing supramolecular compounds: Soluble tetradentate dithioglyoximes for the detection of organic solvents in the gas phase

    SciTech Connect

    Oeztuerk, Z.Z.

    1996-12-31

    Supramolecular structures have been widely investigated for their specific recognition of ions. Recently, they have been also used to detect neutral molecules and especially organic solvents in the gas phase. In this context, the authors developed a new type of supramolecular structures for the recognition of the organic solvent molecules by their selective incorporation to the bulk of thin films. The authors synthesized a series of the soluble tetradentate dithiogloximes which included mono-, tri-, and pentanuclear metal complexes as well as the heteronuclear polymers. Thin layers of these compounds were prepared from their solutions by evaporating their volatile organic solvents. The subsequent exposure of these films to the same organic solvent molecules in air shows drastic transducer signals, i.e., changes in masses and capacitances due to selective interactions of coordinatively and electronically unsaturated metal complexes with gas molecules. The organic solvents investigated in this work are dichloromethane, trichloromethane, tetrachloromethane, trichloroethylene, tetrachloroethylene, acetone, benzene, toluene, n-hexane, n-octane, methanol, ethanol and n-propanol. Sensor responses were monitored with interdigital capacitor (IDC) and quartz crystal oscillators (QCM).

  3. Spectroscopic and Theoretical Study on the Structures and Dynamics of Functional Molecules - Towards AN Understanding of the Molecular Recognition for Encapsulation Complexes

    NASA Astrophysics Data System (ADS)

    Ebata, Takayuki; Kusaka, Ryoji; Inokuchi, Yoshiya; Xantheas, Sotiris S.

    2011-06-01

    Functional molecules, such as crown ethers and calixarenes, can act as hosts for encapsulating guest species through non-covalent interactions. Applications of crown ethers and calixarenes as molecular receptors, metal cation extraction agents, fluoro-ionophores and phase transfer catalytic media have been previously reported in a number of studies in the literature. One of the important aspects of these host/guest molecular assemblies is their selectivity in the encapsulation of guest species. Two important factors that control this selectivity are: (1) the size and the flexibility of the host cavity and (2) the properties of solvent molecules. Molecular complexes formed in supersonic jets provide ideal systems for the selective study of the conformational preference and micro-solvated effects under solvent-controlled conditions. This talk will review our spectroscopic and theoretical studies of the structures of dibenzo-18-crown-6-ether (DB18C6), benzo-18-crown-6-ether (B18C6), calix[4]arene (C4A) and their complexes with guest molecules. We apply laser-induced fluorescence (LIF), resonance enhanced two-photon ionization (R2PI) and UV-UV hole-burning (HB) spectroscopy for obtaining electronic spectra and IR-UV doubleresonance and IR photodissociation (IRPD) spectroscopy for the IR spectra. The electronic and IR spectra are compared with the corresponding results obtained by DFT calculations and high-level first principles electronic structure calculations [MP2 and CCSD(T)]. Based on these joint studies we can elucidate the nature of interactions that control the encapsulation of a guest molecular species as well as how the host can adjust its conformation to accommodate a specific guest, leading to the molecular recognition.

  4. [Strategies to choose scaffold materials for tissue engineering].

    PubMed

    Gao, Qingdong; Zhu, Xulong; Xiang, Junxi; Lü, Yi; Li, Jianhui

    2016-02-01

    Current therapies of organ failure or a wide range of tissue defect are often not ideal. Transplantation is the only effective way for long time survival. But it is hard to meet huge patients demands because of donor shortage, immune rejection and other problems. Tissue engineering could be a potential option. Choosing a suitable scaffold material is an essential part of it. According to different sources, tissue engineering scaffold materials could be divided into three types which are natural and its modified materials, artificial and composite ones. The purpose of tissue engineering scaffold is to repair the tissues or organs damage, so could reach the ideal recovery in its function and structure aspect. Therefore, tissue engineering scaffold should even be as close as much to the original tissue or organs in function and structure. We call it "organic scaffold" and this strategy might be the drastic perfect substitute for the tissues or organs in concern. Optimized organization with each kind scaffold materials could make up for biomimetic structure and function of the tissue or organs. Scaffold material surface modification, optimized preparation procedure and cytosine sustained-release microsphere addition should be considered together. This strategy is expected to open new perspectives for tissue engineering. Multidisciplinary approach including material science, molecular biology, and engineering might find the most ideal tissue engineering scaffold. Using the strategy of drawing on each other strength and optimized organization with each kind scaffold material to prepare a multifunctional biomimetic tissue engineering scaffold might be a good method for choosing tissue engineering scaffold materials. Our research group had differentiated bone marrow mesenchymal stem cells into bile canaliculi like cells. We prepared poly(L-lactic acid)/poly(ε-caprolactone) biliary stent. The scaffold's internal played a part in the long-term release of cytokines which

  5. Nα-terminal acetylation for T cell recognition: molecular basis of MHC class I-restricted nα-acetylpeptide presentation.

    PubMed

    Sun, Mingwei; Liu, Jun; Qi, Jianxun; Tefsen, Boris; Shi, Yi; Yan, Jinghua; Gao, George F

    2014-06-15

    As one of the most common posttranslational modifications (PTMs) of eukaryotic proteins, N(α)-terminal acetylation (Nt-acetylation) generates a class of N(α)-acetylpeptides that are known to be presented by MHC class I at the cell surface. Although such PTM plays a pivotal role in adjusting proteolysis, the molecular basis for the presentation and T cell recognition of N(α)-acetylpeptides remains largely unknown. In this study, we determined a high-resolution crystallographic structure of HLA (HLA)-B*3901 complexed with an N(α)-acetylpeptide derived from natural cellular processing, also in comparison with the unmodified-peptide complex. Unlike the α-amino-free P1 residues of unmodified peptide, of which the α-amino group inserts into pocket A of the Ag-binding groove, the N(α)-linked acetyl of the acetylated P1-Ser protrudes out of the groove for T cell recognition. Moreover, the Nt-acetylation not only alters the conformation of the peptide but also switches the residues in the α1-helix of HLA-B*3901, which may impact the T cell engagement. The thermostability measurements of complexes between N(α)-acetylpeptides and a series of MHC class I molecules derived from different species reveal reduced stability. Our findings provide the insight into the mode of N(α)-acetylpeptide-specific presentation by classical MHC class I molecules and shed light on the potential of acetylepitope-based immune intervene and vaccine development. PMID:24829406

  6. Molecular cloning and characterisation of a pattern recognition protein, lipopolysaccharide and beta-1,3-glucan binding protein (LGBP) from Chinese shrimp Fenneropenaeus chinensis.

    PubMed

    Liu, Fengsong; Li, Fuhua; Dong, Bo; Wang, Xiaomei; Xiang, Jianhai

    2009-03-01

    A pattern recognition protein (PRP), lipopolysaccharide and beta-1,3-glucan binding protein (LGBP) cDNA was cloned from the haemocyte of Chinese shrimp Fenneropenaeus chinensis by the techniques of homology cloning and RACE. Analysis of nucleotide sequence revealed that the full-length cDNA of 1,275 bp has an open reading frame of 1,098 bp encoding a protein of 366 amino acids including a 17 amino acid signal peptide. Sequence comparison of the deduced amino acid sequence of F. chinensis LGBP showed a high identity of 94%, 90%, 87%, 72% and 63% with Penaeus monodon BGBP, Litopenaeus stylirostris LGBP, Marsupenaeu japonicus BGBP, Homarus gammarus BGBP and Pacifastacus leniusculus LGBP, respectively. The calculated molecular mass of the mature protein is 39,857 Da with a deduced pI of 4.39. Two putative integrin binding motifs, RGD (Arg-Gly-Asp) and a potential recognition motif for beta-1,3-linkage of polysaccharides were observed in LGBP sequence. RT-PCR analysis showed that LGBP gene expresses in haemocyte and hepatopancreas only, but not in other tissues. Capillary electrophoresis RT-PCR method was used to quantify the variation of mRNA transcription level during artificial infection with heat-killed Vibrio anguillarum and Staphylococcus aureusin. A significant enhancement of LGBP transcription was appeared at 6 h post-injection in response to bacterial infection. These results have provided useful information to understand the function of LGBP in shrimp. PMID:18163220

  7. Molecular Recognition of 6′-N-5-Hexynoate Kanamyin A and RNA 1×1 Internal Loops Containing CA Mismatches

    PubMed Central

    Tran, Tuan; Disney, Matthew D.

    2011-01-01

    In our previous study to identify the RNA internal loops that bind an aminoglycoside derivative, we determined that 6′-N-5-hexynoate kanamycin A prefers to bind 1×1 nucleotide internal loops containing C•A mismatches. In this present study, the molecular recognition between a variety of RNAs that are mutated around the C•A loop and the ligand was investigated. Studies show that both loop nucleotides and loop closing pairs affect binding affinity. Most interestingly, it was shown that there is a correlation between the thermodynamic stability of the C•A internal loops and ligand affinity. Specifically, C•A loops that had relatively high or low stability bound the ligand most weakly whereas loops with intermediate stability bound the ligand most tightly. In contrast, there is no correlation between the likelihood that a loop forms a C-A+ pair at lower pH and ligand affinity. It was also found that a 1×1 nucleotide C•A loop that bound to the ligand with the highest affinity is identical to the consensus site in RNAs that are edited by adenosine deaminases acting on RNA type 2 (ADAR2). These studies provide a detailed investigation of factors affecting small molecule recognition of internal loops containing C•A mismatches, which are present in a variety of RNAs that cause disease. PMID:21207945

  8. Towards biomimetic scaffolds: anhydrous scaffold fabrication from biodegradable amine-reactive diblock copolymers.

    PubMed

    Hacker, Michael; Tessmar, Jörg; Neubauer, Markus; Blaimer, Andrea; Blunk, Torsten; Göpferich, Achim; Schulz, Michaela B

    2003-11-01

    The development of biomimetic materials and their processing into three-dimensional cell carrying scaffolds is one promising tissue engineering strategy to improve cell adhesion, growth and differentiation on polymeric constructs developing mature and viable tissue. This study was concerned with the fabrication of scaffolds made from amine-reactive diblock copolymers, N-succinimidyl tartrate monoamine poly(ethylene glycol)-block-poly(D,L-lactic acid), which are able to suppress unspecific protein adsorption and to covalently bind proteins or peptides. An appropriate technique for their processing had to be both anhydrous, to avoid hydrolysis of the active ester, and suitable for the generation of interconnected porous structures. Attempts to fabricate scaffolds utilizing hard paraffin microparticles as hexane-extractable porogens failed. Consequently, a technique was developed involving lipid microparticles, which served as biocompatible porogens on which the scaffold forming polymer was precipitated in the porogen extraction media (n-hexane). Porogen melting during the extraction and polymer precipitation step led to an interconnected network of pores. Suitable lipid mixtures and their melting points, extraction conditions (temperature and time) and a low-toxic polymer solvent system were determined for their use in processing diblock copolymers of different molecular weights (22 and 42 kDa) into highly porous off-the-shelf cell carriers ready for easy surface modification towards biomimetic scaffolds. Insulin was employed to demonstrate the principal of instant protein coupling to a prefabricated scaffold. PMID:12922156

  9. Tetranitro-oxacalix[4]crown-Based Host-Guest Recognition Motif and a Related [2]Rotaxane-Based Molecular Switch.

    PubMed

    Liu, Hua; Hu, Wen-Jing; Liu, Yahu A; Li, Jiu-Sheng; Jiang, Biao; Wen, Ke

    2016-08-01

    Different from so far reported oxacalix[4]crown-based host-guest motifs in which oxacalix[4]crowns act only as hydrogen bond acceptors, a [2]pseudorotaxane-type tetranitro-oxacalix[4]crown/urea host-guest recognition motif was developed in which tetranitro-oxacalix[4]crown played a role as both a hydrogen bond donor and an acceptor to stabilize the resulting supramolecular complex. Furthermore, on the basis of a [2]pseudorotaxane complex formed from a tetranitro-oxacalix[4]crown and an axle containing a secondary ammonium ion and a urea group, a [2]rotaxane-based molecular switch was created, in which the oxacalix[4]crown wheel was able to reversibly translocate between the secondary ammonium binding site and the urea binding site of the axle under acid-base stimulation. PMID:27384815

  10. Macroscopic observations of molecular recognition: discrimination of the substituted position on the naphthyl group by polyacrylamide gel modified with β-cyclodextrin.

    PubMed

    Zheng, Yongtai; Hashidzume, Akihito; Takashima, Yoshinori; Yamaguchi, Hiroyasu; Harada, Akira

    2011-11-15

    Macroscopic molecular recognition observations were realized using polyacrylamide-based gels modified with α-cyclodextrin (α-CD), β-cyclodextrin (β-CD), 1-naphthylmethyl (1Np), and 2-naphthylmethyl (2Np) moieties, which are denoted as αCD(x)-gel, βCD(x)-gel, 1Np(y)-gel, and 2Np(y)-gel, where x and y indicate the mol % of CD and Np moieties, respectively. The αCD(5)-gel did not adhere to either the 1Np(5)-gel or 2Np(5)-gel, whereas the βCD(5)-gel interacted with both to form alternating or checkered assemblies. Although the difference in the association constants of β-CD for the model polymers was small, the βCD(x)-gel successfully discriminated between 1Np(y)-gel and 2Np(y)-gel at the appropriate x and y. PMID:21978319

  11. Identification of anticancer agents based on the thieno[2,3-b]pyridine and 1H-pyrazole molecular scaffolds.

    PubMed

    Eurtivong, Chatchakorn; Reynisdóttir, Inga; Kuczma, Stephanie; Furkert, Daniel P; Brimble, Margaret A; Reynisson, Jóhannes

    2016-08-15

    Structural similarity search of commercially available analogues of thieno[2,3-b]pyridine and 1H-pyrazole derivatives, known anticancer agents, resulted in 717 hits. These were docked into the phosphoinositide specific-phospholipase C (PLC) binding pocket, the putative target of the compounds, to further focus the selection. Thirteen derivatives of the thieno[2,3-b]pyridines were identified and tested against the NCI60 panel of human tumour cell lines. The most active derivative 1 was most potent against the MDA-MB-435 melanoma cell line with GI50 at 30nM. Also, it was found that a piperidine moiety is tolerated on the thieno[2,3-b]pyridine scaffold with GI50=296nM (MDA-MB-435) for derivative 10 considerably expanding the structure activity relationship for the series. For the 1H-pyrazoles four derivatives were identified using the in silico approach and additionally ten were synthesised with various substituents on the phenyl moiety to extend the structural activity relationship but only modest anticancer activity was found. PMID:27288184

  12. Exact approaches for scaffolding

    PubMed Central

    2015-01-01

    This paper presents new structural and algorithmic results around the scaffolding problem, which occurs prominently in next generation sequencing. The problem can be formalized as an optimization problem on a special graph, the "scaffold graph". We prove that the problem is polynomial if this graph is a tree by providing a dynamic programming algorithm for this case. This algorithm serves as a basis to deduce an exact algorithm for general graphs using a tree decomposition of the input. We explore other structural parameters, proving a linear-size problem kernel with respect to the size of a feedback-edge set on a restricted version of Scaffolding. Finally, we examine some parameters of scaffold graphs, which are based on real-world genomes, revealing that the feedback edge set is significantly smaller than the input size. PMID:26451725

  13. Use of UV-vis-NIR spectroscopy to monitor label-free interaction between molecular recognition elements and erythropoietin on a gold-coated polycarbonate platform.

    PubMed

    Citartan, Marimuthu; Gopinath, Subash C B; Tominaga, Junji; Chen, Yeng; Tang, Thean-Hock

    2014-08-01

    Label-free-based detection is pivotal for real-time monitoring of biomolecular interactions and to eliminate the need for labeling with tags that can occupy important binding sites of biomolecules. One simplest form of label-free-based detection is ultraviolet-visible-near-infrared (UV-vis-NIR) spectroscopy, which measure changes in reflectivity as a means to monitor immobilization and interaction of biomolecules with their corresponding partners. In biosensor development, the platform used for the biomolecular interaction should be suitable for different molecular recognition elements. In this study, gold (Au)-coated polycarbonate was used as a platform and as a proof-of-concept, erythropoietin (EPO), a doping substance widely abused by the athletes was used as the target. The interaction of EPO with its corresponding molecular recognition elements (anti-EPO monoclonal antibody and anti-EPO DNA aptamer) is monitored by UV-vis-NIR spectroscopy. Prior to this, to show that UV-vis-NIR spectroscopy is a suitable method for measuring biomolecular interaction, the interaction between biotin and streptavidin was demonstrated via this strategy and reflectivity of this interaction decreased by 25%. Subsequent to this, interaction of the EPO with anti-EPO monoclonal antibody and anti-EPO DNA aptamer resulted in the decrease of reflectivity by 5% and 10%, respectively. The results indicated that Au-coated polycarbonate could be an ideal biosensor platform for monitoring biomolecular interactions using UV-vis-NIR spectroscopy. A smaller version of the Au-coated polycarbonate substrates can be derived from the recent set-up, to be applied towards detecting EPO abuse among atheletes. PMID:24881539

  14. Molecular basis of the recognition of the ap65-1 gene transcription promoter elements by a Myb protein from the protozoan parasite Trichomonas vaginalis.

    PubMed

    Jiang, Ingjye; Tsai, Chen-Kun; Chen, Sheng-Chia; Wang, Szu-Huan; Amiraslanov, Imamaddin; Chang, Chi-Fon; Wu, Wen-Jin; Tai, Jung-Hsiang; Liaw, Yen-Chywan; Huang, Tai-Huang

    2011-11-01

    Iron-inducible transcription of the ap65-1 gene in Trichomonas vaginalis involves at least three Myb-like transcriptional factors (tvMyb1, tvMyb2 and tvMyb3) that differentially bind to two closely spaced promoter sites, MRE-1/MRE-2r and MRE-2f. Here, we defined a fragment of tvMyb2 comprising residues 40-156 (tvMyb2₄₀₋₁₅₆) as the minimum structural unit that retains near full binding affinity with the promoter DNAs. Like c-Myb in vertebrates, the DNA-free tvMyb2₄₀₋₁₅₆ has a flexible and open conformation. Upon binding to the promoter DNA elements, tvMyb2₄₀₋₁₅₆ undergoes significant conformational re-arrangement and structure stabilization. Crystal structures of tvMyb2₄₀₋₁₅₆ in complex with promoter element-containing DNA oligomers showed that 5'-a/gACGAT-3' is the specific base sequence recognized by tvMyb2₄₀₋₁₅₆, which does not fully conform to that of the Myb binding site sequence. Furthermore, Lys⁴⁹, which is upstream of the R2 motif (amino acids 52-102) also participates in specific DNA sequence recognition. Intriguingly, tvMyb2₄₀₋₁₅₆ binds to the promoter elements in an orientation opposite to that proposed in the HADDOCK model of the tvMyb1₃₅₋₁₄₁/MRE-1-MRE-2r complex. These results shed new light on understanding the molecular mechanism of Myb-DNA recognition and provide a framework to study the molecular basis of transcriptional regulation of myriad Mybs in T. vaginalis. PMID:21771861

  15. A malonitrile-functionalized metal-organic framework for hydrogen sulfide detection and selective amino acid molecular recognition

    PubMed Central

    Li, Haiwei; Feng, Xiao; Guo, Yuexin; Chen, Didi; Li, Rui; Ren, Xiaoqian; Jiang, Xin; Dong, Yuping; Wang, Bo

    2014-01-01

    A novel porous polymeric fluorescence probe, MN-ZIF-90, has been designed and synthesized for quantitative hydrogen sulfide (H2S) fluorescent detection and highly selective amino acid recognition. This distinct crystalline structure, derived from rational design and malonitrile functionalization, can trigger significant enhancement of its fluorescent intensity when exposed to H2S or cysteine molecules. Indeed this new metal-organic framework (MOF) structure shows high selectivity of biothiols over other amino acids and exhibits favorable stability. Moreover, in vitro viability assays on HeLa cells show low cytotoxicity of MN-ZIF-90 and its imaging contrast efficiency is further demonstrated by fluorescence microscopy studies. This facile yet powerful strategy also offers great potential of using open-framework materials (i.e. MOFs) as the novel platform for sensing and other biological applications. PMID:24621614

  16. Scaffolds in Tendon Tissue Engineering

    PubMed Central

    Longo, Umile Giuseppe; Lamberti, Alfredo; Petrillo, Stefano; Maffulli, Nicola; Denaro, Vincenzo

    2012-01-01

    Tissue engineering techniques using novel scaffold materials offer potential alternatives for managing tendon disorders. Tissue engineering strategies to improve tendon repair healing include the use of scaffolds, growth factors, cell seeding, or a combination of these approaches. Scaffolds have been the most common strategy investigated to date. Available scaffolds for tendon repair include both biological scaffolds, obtained from mammalian tissues, and synthetic scaffolds, manufactured from chemical compounds. Preliminary studies support the idea that scaffolds can provide an alternative for tendon augmentation with an enormous therapeutic potential. However, available data are lacking to allow definitive conclusion on the use of scaffolds for tendon augmentation. We review the current basic science and clinical understanding in the field of scaffolds and tissue engineering for tendon repair. PMID:22190961

  17. High-resolution single-molecule recognition imaging of the molecular details of ricin-aptamer interaction

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The molecular details of DNA aptamer-ricin interactions were investigated. The toxic protein ricin molecules were immobilized on Au(111) surface using N-hydroxysuccinimide (NHS) ester to specifically react with lysine residues located on the ricin B chains. A single ricin molecule was visualized in ...

  18. RECOGNITION OF PYRENE USING MOLECULARLY-IMPRINTED ELECTROCHEMICALLY-DEPOSITED POLY (2-MERCAPTOBENZIMIDAZOLE) OR POLY(RESORCINOL) ON GOLD ELECTRODES

    EPA Science Inventory

    The feasibility of using thiol chemistry to form molecularly imprinted polymer-coated gold electrodes to measure pyrene is reported. For the first approach, poly(2-mercaptoimidazole) (2-MBI) was electrochemically deposited on gold electrodes in the presence or absence of the tem...

  19. Molecularly imprinted optosensing material based on hydrophobic CdSe quantum dots via a reverse microemulsion for specific recognition of ractopamine.

    PubMed

    Liu, Huilin; Fang, Guozhen; Wang, Shuo

    2014-05-15

    A novel molecularly imprinted polymer (MIP) based on hydrophobic CdSe quantum dots (QDs) was synthesized using a one-pot room-temperature reverse microemulsion polymerization, and this polymer was applied as a molecular recognition element to construct a ractopamine (RAC) optosensor. Here, hydrophobic CdSe QDs were first introduced to the hydrophilic analyte-imprinted polymer for highly selective and sensitive detection of RAC via the change in fluorescence intensity, because of the high-quality hydrophobic QDs with high quantum yield, sharp photoluminescence spectra and chemical and fluorescent stability. Under optimal conditions, the relative fluorescence intensity of MIP based on hydrophobic QDs decreased linearly with the increasing concentration of RAC in the range of 1.21 × 10(-9) -3.03 × 10(-6)mol L(-1) with a detection limit of 7.57 × 10(-10)mol L(-1), and the precision for five replicate detections of 1.51 × 10(-8)mol L(-1) RAC was 2.09% (relative standard deviation). The proposed method was successfully applied for the determination of trace RAC in pork samples, with good recoveries ranging from 82.79% to 97.23%. PMID:24370883

  20. Molecularly imprinted electrochemical sensing interface based on in-situ-polymerization of amino-functionalized ionic liquid for specific recognition of bovine serum albumin.

    PubMed

    Wang, Yanying; Han, Miao; Liu, Guishen; Hou, Xiaodong; Huang, Yina; Wu, Kangbing; Li, Chunya

    2015-12-15

    A molecularly imprinted polymer film was in situ polymerized on a carboxyl functionalized multi-walled carbon nanotubes modified glassy carbon electrode surface under room temperature. This technique provides a promising imprinting approach for protein in an aqueous solution using 3-(3-aminopropyl)-1-vinylimidazolium tetrafluoroborate ionic liquid as functional monomer, N, N'-methylenebisacrylamide as crossing linker, ammonium persulfate and N,N,N',N'-tetramethylethylenediamine as initiator, and bovine serum albumin (BSA) as template. The molecularly imprinted polymerized ionic liquid film shows enhanced accessibility, high specificity and sensitivity towards BSA. Electrochemical sensing performance of the imprinted sensor was thoroughly investigated using K3Fe[CN]6/K4Fe[CN]6 as electroactive probes. Under optimal conditions, the current difference before and after specific recognition of BSA was found linearly related to its concentration in the range from 1.50×10(-9) to 1.50×10(-6) mol L(-1). The detection limit was calculated to be 3.91×10(-10) mol L(-1) (S/N=3). The practical application of the imprinted sensor was demonstrated by determining BSA in liquid milk samples. PMID:26232004

  1. Molecular Basis for the Selectivity and Specificity of Ligand Recognition by the Family 16 Carbohydrate-binding Modules from Thermoanaerobacterium polysaccharolyticum ManA

    SciTech Connect

    Bae, Brian; Ohene-Adjei, Samuel; Kocherginskaya, Svetlana; Mackie, Roderick I.; Spies, M. Ashley; Cann, Isaac K.O.; Nair, Satish K.

    2008-06-06

    Enzymes that hydrolyze complex polysaccharides into simple sugars are modular in architecture and consist of single or multiple catalytic domains fused to targeting modules called carbohydrate-binding modules (CBMs). CBMs bind to their ligands with high affinity and increase the efficiency of the catalytic components by targeting the enzymes to its substrate. Here we utilized a multidisciplinary approach to characterize each of the two family 16 carbohydrate-binding domain components of the highly active mannanase from the thermophile Thermoanaerobacterium polysaccharolyticum. These represent the first crystal structures of family 16 CBMs. Calorimetric analysis showed that although these CBMs demonstrate high specificity toward {beta}-1,4-linked sugars, they can engage both cello- and mannopolysaccharides. To elucidate the molecular basis for this specificity and selectivity, we have determined high resolution crystal structures of each of the two CBMs, as well as of binary complexes of CBM16-1 bound to either mannopentaose or cellopentaose. These results provide detailed molecular insights into ligand recognition and yield a framework for rational engineering experiments designed to expand the natural repertoire of these targeting modules.

  2. Selective Recognition of d-Aldohexoses in Water by Boronic Acid-Functionalized, Molecularly Imprinted Cross-Linked Micelles.

    PubMed

    Awino, Joseph K; Gunasekara, Roshan W; Zhao, Yan

    2016-08-10

    Molecular imprinting within cross-linked micelles using 4-vinylphenylboronate derivatives of carbohydrates provided water-soluble nanoparticle receptors selective for the carbohydrate templates. Complete differentiation of d-aldohexoses could be achieved by these receptors if a single inversion of hydroxyl occurred at C2 or C4 of the sugar or if two or more inversions took place. Glycosides with a hydrophobic aglycan displayed stronger binding due to increased hydrophobic interactions. PMID:27442012

  3. Structure of the WD40 domain of SCAP from fission yeast reveals the molecular basis for SREBP recognition.

    PubMed

    Gong, Xin; Li, Jingxian; Shao, Wei; Wu, Jianping; Qian, Hongwu; Ren, Ruobing; Espenshade, Peter; Yan, Nieng

    2015-04-01

    The sterol regulatory element-binding protein (SREBP) and SREBP cleavage-activating protein (SCAP) are central players in the SREBP pathway, which control the cellular lipid homeostasis. SCAP binds to SREBP through their carboxyl (C) domains and escorts SREBP from the endoplasmic reticulum to the Golgi upon sterol depletion. A conserved pathway, with the homologues of SREBP and SCAP being Sre1 and Scp1, was identified in fission yeast Schizosaccharomyces pombe. Here we report the in vitro reconstitution of the complex between the C domains of Sre1 and Scp1 as well as the crystal structure of the WD40 domain of Scp1 at 2.1 Å resolution. The structure reveals an eight-bladed β-propeller that exhibits several distinctive features from a canonical WD40 repeat domain. Structural and biochemical characterization led to the identification of two Scp1 elements that are involved in Sre1 recognition, an Arg/Lys-enriched surface patch on the top face of the WD40 propeller and a 30-residue C-terminal tail. The structural and biochemical findings were corroborated by in vivo examinations. These studies serve as a framework for the mechanistic understanding and further functional characterization of the SREBP and SCAP proteins in fission yeast and higher organisms. PMID:25771684

  4. Structure of the WD40 domain of SCAP from fission yeast reveals the molecular basis for SREBP recognition

    PubMed Central

    Gong, Xin; Li, Jingxian; Shao, Wei; Wu, Jianping; Qian, Hongwu; Ren, Ruobing; Espenshade, Peter; Yan, Nieng

    2015-01-01

    The sterol regulatory element-binding protein (SREBP) and SREBP cleavage-activating protein (SCAP) are central players in the SREBP pathway, which control the cellular lipid homeostasis. SCAP binds to SREBP through their carboxyl (C) domains and escorts SREBP from the endoplasmic reticulum to the Golgi upon sterol depletion. A conserved pathway, with the homologues of SREBP and SCAP being Sre1 and Scp1, was identified in fission yeast Schizosaccharomyces pombe. Here we report the in vitro reconstitution of the complex between the C domains of Sre1 and Scp1 as well as the crystal structure of the WD40 domain of Scp1 at 2.1 Å resolution. The structure reveals an eight-bladed β-propeller that exhibits several distinctive features from a canonical WD40 repeat domain. Structural and biochemical characterization led to the identification of two Scp1 elements that are involved in Sre1 recognition, an Arg/Lys-enriched surface patch on the top face of the WD40 propeller and a 30-residue C-terminal tail. The structural and biochemical findings were corroborated by in vivo examinations. These studies serve as a framework for the mechanistic understanding and further functional characterization of the SREBP and SCAP proteins in fission yeast and higher organisms. PMID:25771684

  5. Molecular Mechanisms of Viral and Host Cell Substrate Recognition by Hepatitis C Virus NS3/4A Protease

    SciTech Connect

    Romano, Keith P.; Laine, Jennifer M.; Deveau, Laura M.; Cao, Hong; Massi, Francesca; Schiffer, Celia A.

    2011-08-16

    Hepatitis C NS3/4A protease is a prime therapeutic target that is responsible for cleaving the viral polyprotein at junctions 3-4A, 4A4B, 4B5A, and 5A5B and two host cell adaptor proteins of the innate immune response, TRIF and MAVS. In this study, NS3/4A crystal structures of both host cell cleavage sites were determined and compared to the crystal structures of viral substrates. Two distinct protease conformations were observed and correlated with substrate specificity: (i) 3-4A, 4A4B, 5A5B, and MAVS, which are processed more efficiently by the protease, form extensive electrostatic networks when in complex with the protease, and (ii) TRIF and 4B5A, which contain polyproline motifs in their full-length sequences, do not form electrostatic networks in their crystal complexes. These findings provide mechanistic insights into NS3/4A substrate recognition, which may assist in a more rational approach to inhibitor design in the face of the rapid acquisition of resistance.

  6. Binding of fluorescent acridine dyes acridine orange and 9-aminoacridine to hemoglobin: Elucidation of their molecular recognition by spectroscopy, calorimetry and molecular modeling techniques.

    PubMed

    Chatterjee, Sabyasachi; Kumar, Gopinatha Suresh

    2016-06-01

    The molecular interaction between hemoglobin (HHb), the major human heme protein, and the acridine dyes acridine orange (AO) and 9-aminoacridine (9AA) was studied by various spectroscopic, calorimetric and molecular modeling techniques. The dyes formed stable ground state complex with HHb as revealed from spectroscopic data. Temperature dependent fluorescence data showed the strength of the dye-protein complexation to be inversely proportional to temperature and the fluorescence quenching was static in nature. The binding-induced conformational change in the protein was investigated using circular dichroism, synchronous fluorescence, 3D fluorescence and FTIR spectroscopy results. Circular dichroism data also quantified the α-helicity change in hemoglobin due to the binding of acridine dyes. Calorimetric studies revealed the binding to be endothermic in nature for both AO and 9AA, though the latter had higher affinity, and this was also observed from spectroscopic data. The binding of both dyes was entropy driven. pH dependent fluorescence studies revealed the existence of electrostatic interaction between the protein and dye molecules. Molecular modeling studies specified the binding site and the non-covalent interactions involved in the association. Overall, the results revealed that a small change in the acridine chromophore leads to remarkable alteration in the structural and thermodynamic aspects of binding to HHb. PMID:27077554

  7. Synthesis of carbohydrate-scaffolded thymine glycoconjugates to organize multivalency.

    PubMed

    Ciuk, Anna K; Lindhorst, Thisbe K

    2015-01-01

    Multivalency effects are essential in carbohydrate recognition processes as occurring on the cell surface. Thus many synthetic multivalent glycoconjugates have been developed as important tools for glycobiological research. We are expanding this collection of molecules by the introduction of carbohydrate-scaffolded divalent glycothymine derivatives that can be intramolecularily dimerized by [2 + 2] photocycloaddition. Thus, thymine functions as a control element that allows to restrict the conformational flexibility of the scaffolded sugar ligands and thus to "organize" multivalency. With this work we add a parameter to multivalency studies additional to valency. PMID:26124869

  8. Synthesis of carbohydrate-scaffolded thymine glycoconjugates to organize multivalency

    PubMed Central

    Ciuk, Anna K

    2015-01-01

    Summary Multivalency effects are essential in carbohydrate recognition processes as occurring on the cell surface. Thus many synthetic multivalent glycoconjugates have been developed as important tools for glycobiological research. We are expanding this collection of molecules by the introduction of carbohydrate-scaffolded divalent glycothymine derivatives that can be intramolecularily dimerized by [2 + 2] photocycloaddition. Thus, thymine functions as a control element that allows to restrict the conformational flexibility of the scaffolded sugar ligands and thus to “organize” multivalency. With this work we add a parameter to multivalency studies additional to valency. PMID:26124869

  9. Insight into the Intermolecular Recognition Mechanism between Keap1 and IKKβ Combining Homology Modelling, Protein-Protein Docking, Molecular Dynamics Simulations and Virtual Alanine Mutation

    PubMed Central

    Jiang, Zheng-Yu; Chu, Hong-Xi; Xi, Mei-Yang; Yang, Ting-Ting; Jia, Jian-Min; Huang, Jing-Jie; Guo, Xiao-Ke; Zhang, Xiao-Jin; You, Qi-Dong; Sun, Hao-Peng

    2013-01-01

    Degradation of certain proteins through the ubiquitin-proteasome pathway is a common strategy taken by the key modulators responsible for stress responses. Kelch-like ECH-associated protein-1(Keap1), a substrate adaptor component of the Cullin3 (Cul3)-based ubiquitin E3 ligase complex, mediates the ubiquitination of two key modulators, NF-E2-related factor 2 (Nrf2) and IκB kinase β (IKKβ), which are involved in the redox control of gene transcription. However, compared to the Keap1-Nrf2 protein-protein interaction (PPI), the intermolecular recognition mechanism of Keap1 and IKKβ has been poorly investigated. In order to explore the binding pattern between Keap1 and IKKβ, the PPI model of Keap1 and IKKβ was investigated. The structure of human IKKβ was constructed by means of the homology modeling method and using reported crystal structure of Xenopus laevis IKKβ as the template. A protein-protein docking method was applied to develop the Keap1-IKKβ complex model. After the refinement and visual analysis of docked proteins, the chosen pose was further optimized through molecular dynamics simulations. The resulting structure was utilized to conduct the virtual alanine mutation for the exploration of hot-spots significant for the intermolecular interaction. Overall, our results provided structural insights into the PPI model of Keap1-IKKβ and suggest that the substrate specificity of Keap1 depend on the interaction with the key tyrosines, namely Tyr525, Tyr574 and Tyr334. The study presented in the current project may be useful to design molecules that selectively modulate Keap1. The selective recognition mechanism of Keap1 with IKKβ or Nrf2 will be helpful to further know the crosstalk between NF-κB and Nrf2 signaling. PMID:24066166

  10. Clean synthesis of molecular recognition polymeric materials with chiral sensing capability using supercritical fluid technology. Application as HPLC stationary phases.

    PubMed

    da Silva, Mara Soares; Vão, Eva R; Temtem, Márcio; Mafra, Luís; Caldeira, Jorge; Aguiar-Ricardo, Ana; Casimiro, Teresa

    2010-03-15

    Molecularly imprinted polymers (MIPs) of poly(ethylene glycol dimethacrylate) and poly(N-isopropylacrylamide-co-ethylene glycol dimethacrylate) were synthesized for the first time in supercritical carbon dioxide (scCO(2)), using Boc-L-tryptophan as template. Supercritical fluid technology provides a clean and one-step synthetic route for the preparation of affinity polymeric materials with sensing capability for specific molecules. The polymeric materials were tested as stationary HPLC phases for the enantiomeric separation of L- and D-tryptophan. HPLC results prove that the synthesized MIPs are able to recognize the template molecule towards its enantiomer which opens up potential applications in chromatographic chiral separation. PMID:20096557

  11. Molecularly imprinted macroporous monoliths for solid-phase extraction: Effect of pore size and column length on recognition properties.

    PubMed

    Vlakh, E G; Stepanova, M A; Korneeva, Yu M; Tennikova, T B

    2016-09-01

    The series of macroporous monolithic molecularly imprinted monoliths differed by pore size, column length (volume) and amount of template used for imprinting was synthesized using methacrylic acid and glycerol dimethacrylate as co-monomers and antibiotic ciprofloxacin as a template. The prepared monoliths were characterized regarding to their permeability, pore size, porosity, and resistance to the flow of a mobile phase. The surface morphology was also analyzed. The slight dependence of imprinting factor on flow rate, as well as its independence on pore size of macroporous molecularly imprinted monolithic media was observed. The column obtained at different conditions exhibited different affinity of ciprofloxacin to the imprinted sites that was characterized with Kdiss values in the range of 10(-5)-10(-4)M. The solid-phase extraction of ciprofloxacin from such biological liquids as human blood serum, human urine and cow milk serum was performed using the developed monolithic columns. In all cases, the extraction was found to be 95.0-98.6%. Additionally, the comparison of extraction of three fluoroqinolone analogues, e.g. ciprofloxacin, levofloxacin and moxifloxacin, from human blood plasma was carried out. Contrary to ciprofloxacin extracted with more than 95%, this parameter did not exceed 40% for its analogues. PMID:27433985

  12. Selective recognition and discrimination of water-soluble azo dyes by a seven-channel molecularly imprinted polymer sensor array.

    PubMed

    Long, Zerong; Lu, Yi; Zhang, Mingliang; Qiu, Hongdeng

    2014-10-01

    A seven-channel molecularly imprinted polymer sensor array was prepared and characterized by scanning electron microscopy, Fourier transform infrared spectroscopy, UV-Vis spectroscopy, and nitrogen physisorption studies. The results revealed that the imprinted polymers have distinct-binding affinities from those of structurally similar azo dyes. Analysis of the UV-Vis spectral response patterns of the seven dye analytes against the imprinted polymer array suggested that the different selectivity patterns of the array were closely connected to the imprinting process. To evaluate the effectiveness of the array format, the binding of a series of analytes was individually measured for each of the seven polymers, made with different templates (including one control polymer synthesized without the use of a template). The response patterns of the array to the selected azo dyes were processed by canonical discriminant analysis. The results showed that the molecularly imprinted array was able to discriminate each analyte with 100% accuracy. Moreover, the azo dyes in two real samples, spiked chrysoidin in smoked bean curd extract and Fanta lime soda (containing tartrazine), were successfully classified by the array. PMID:25099151

  13. Biomimetic Scaffolds for Osteogenesis

    PubMed Central

    Yuan, Nance; Rezzadeh, Kameron S.; Lee, Justine C.

    2015-01-01

    Skeletal regenerative medicine emerged as a field of investigation to address large osseous deficiencies secondary to congenital, traumatic, and post-oncologic conditions. Although autologous bone grafts have been the gold standard for reconstruction of skeletal defects, donor site morbidity remains a significant limitation. To address these limitations, contemporary bone tissue engineering research aims to target delivery of osteogenic cells and growth factors in a defined three dimensional space using scaffolding material. Using bone as a template, biomimetic strategies in scaffold engineering unite organic and inorganic components in an optimal configuration to both support osteoinduction as well as osteoconduction. This article reviews the various structural and functional considerations behind the development of effective biomimetic scaffolds for osteogenesis and highlights strategies for enhancing osteogenesis. PMID:26413557

  14. Hyaluronan production and chondrogenic properties of primary human chondrocyte on gelatin based hematostatic spongostan scaffold

    PubMed Central

    2012-01-01

    Background Autologous chondrocyte transplantation is a promising technique for treatment of cartilage defects. Three dimensional chondrocyte cultures on a scaffold are widely used to retain the chondrogenic phenotype. Using a biodegradable gelatin scaffold is one option for the cell delivery system, but molecular and histological studies of the method have not yet been done. Methods We evaluated the chondrogenic property of the primary human chondrocyte on a gelatin scaffold as compared to a collagen scaffold over a period of 21 days. We examined the production of glycosaminoglycan by quantitative and histological analysis. Gene expression of cartilage-associated molecules was assessed by quantitative RT-PCR. Results The gelatin scaffold showed the ability to promote chondrocyte expansion, chondrogenic phenotype retention at molecular and mRNA levels. Conclusions This scaffold is thus suitable for use as an in vitro model for chondrocyte 3D culture. PMID:23253362

  15. Understanding the recognition mechanisms of Zα domain of human editing enzyme ADAR1 (hZα(ADAR1)) and various Z-DNAs from molecular dynamics simulation.

    PubMed

    Wang, Qianqian; Li, Lanlan; Wang, Xiaoting; Liu, Huanxiang; Yao, Xiaojun

    2014-11-01

    The Z-DNA-binding domain of human double-stranded RNA adenosine deaminase I (hZαADAR1) can specifically recognize the left-handed Z-DNA which preferentially occurs at alternating purine-pyrimidine repeats, especially the CG-repeats. The interactions of hZαADAR1 and Z-DNAs in different sequence contexts can affect many important biological functions including gene regulation and chromatin remodeling. Therefore it is of great necessity to fully understand their recognition mechanisms. However, most existing studies are aimed at the standard CG-repeat Z-DNA rather than the non-CG-repeats, and whether the molecular basis of hZαADAR1 binding to various Z-DNAs are identical or not is still unclear on the atomic level. Here, based on the recently determined crystal structures of three representative non-CG-repeat Z-DNAs (d(CACGTG)2, d(CGTACG)2 and d(CGGCCG)2) in complex with hZαADAR1, 40 ns molecular dynamics simulation together with binding free energy calculation were performed for each system. For comparison, the standard CG-repeat Z-DNA (d(CGCGCG)2) complexed with hZαADAR1 was also simulated. The consistent results demonstrate that nonpolar interaction is the driving force during the protein-DNA binding process, and that polar interaction mainly from helix α3 also provides important contributions. Five common hot-spot residues were identified, namely Lys169, Lys170, Asn173, Arg174 and Tyr177. Hydrogen bond analysis coupled with surface charge distribution further reveal the interfacial information between hZαADAR1 and Z-DNA in detail. All of the analysis illustrate that four complexes share the common key features and the similar binding modes irrespective of Z-DNA sequences, suggesting that Z-DNA recognition by hZαADAR1 is conformation-specific rather than sequence-specific. Additionally, by analyzing the conformational changes of hZαADAR1, we found that the binding of Z-DNA could effectively stabilize hZαADAR1 protein. Our study can provide some valuable

  16. Molecularly imprinted poly(N-vinyl imidazole) based polymers grafted onto nonwoven fabrics for recognition/removal of phloretic acid

    NASA Astrophysics Data System (ADS)

    Llorina Rañada, Ma.; Akbulut, Meshude; Abad, Lucille; Güven, Olgun

    2014-01-01

    A solution of N-vinyl imidazole (VIm), ethylene glycol dimethylacrylate (EGDMA), and phloretic acid (p-hydroxyphenylpropionic acid, HPPA) as functional monomer, crosslinker and template, respectively, were used to graft molecularly imprinted polymer (MIP) onto polyethylene/polypropylene (PE/PP) nonwoven fabric via gamma radiation at room temperature. Control grafted films were also synthesized using the same procedure in the absence of HPPA. Binding performance of the MIP grafts was investigated for different template molecule concentrations and contact time. An imprinting factor for the sample prepared at 5 kGy dose was determined as 2.41 for 50 ppm HPPA solution for 3-h incubation. MIP graft layers were investigated by positron annihilation lifetime spectroscopy (PALS) as well as SEM.

  17. Equilibrium shift in solution: molecular shape recognition and precipitation of a synthetic double helix using helicene-grafted silica nanoparticles.

    PubMed

    Miyagawa, Masamichi; Ichinose, Wataru; Yamaguchi, Masahiko

    2014-01-27

    Chiral silica nanoparticles (70 nm) grafted with (P)-helicene recognized the molecular shape of double helix and random coil (P)-ethynylhelicene oligomers in solution. A mixture of the (P)-nanoparticles and double helix precipitated much faster than a mixture of the (P)-nanoparticles and random coil, and the precipitate contained only the double helix. The mixture of the (P)-nanoparticles and (P)-ethynylhelicene pentamer reversibly dispersed in trifluoromethylbenzene upon heating at 70 °C and precipitated upon cooling at 25 °C. When a 10:90 equilibrium mixture of the double helix and random coil in solution was treated with the (P)-nanoparticles, the double helix was precipitated in 53% yield and was accompanied by equilibrium shift. PMID:24382663

  18. Silks as scaffolds for skin reconstruction.

    PubMed

    Reimers, Kerstin; Liebsch, Christina; Radtke, Christine; Kuhbier, Jörn W; Vogt, Peter M

    2015-11-01

    In this short review, we describe the use of high molecular weight proteins produced in the glands of several arthropods-commonly called silks-for the purpose to enhance human skin wound healing. To this end an extensive literature search has been performed, the publications have been categorized concerning silk preparation and application and summarized accordingly: Scaffolds to promote wound healing were prepared by processing the silks in different ways including solubilization of the protein fibers followed by casting or electrospinning. The silk scaffolds were additionally modified by coating or blending with the intention of further functionalization. In several approaches, the scaffolds were also vitalized with skin cells or stem cells. In vitro and in vivo models were implied to test for safety and efficiency. We conclude that silk scaffolds are characterized by an advantageous biocompatibility as well as an impressive versatility rendering them ideally suited for application in wounds. Nevertheless, further investigation is needed to exploit the full capacity of silk in different wound models and to achieve clinical transfer in time. PMID:25995140

  19. Engineered Biopolymeric Scaffolds for Chronic Wound Healing

    PubMed Central

    Dickinson, Laura E.; Gerecht, Sharon

    2016-01-01

    Skin regeneration requires the coordinated integration of concomitant biological and molecular events in the extracellular wound environment during overlapping phases of inflammation, proliferation, and matrix remodeling. This process is highly efficient during normal wound healing. However, chronic wounds fail to progress through the ordered and reparative wound healing process and are unable to heal, requiring long-term treatment at high costs. There are many advanced skin substitutes, which mostly comprise bioactive dressings containing mammalian derived matrix components, and/or human cells, in clinical use. However, it is presently hypothesized that no treatment significantly outperforms the others. To address this unmet challenge, recent research has focused on developing innovative acellular biopolymeric scaffolds as more efficacious wound healing therapies. These biomaterial-based skin substitutes are precisely engineered and fine-tuned to recapitulate aspects of the wound healing milieu and target specific events in the wound healing cascade to facilitate complete skin repair with restored function and tissue integrity. This mini-review will provide a brief overview of chronic wound healing and current skin substitute treatment strategies while focusing on recent engineering approaches that regenerate skin using synthetic, biopolymeric scaffolds. We discuss key polymeric scaffold design criteria, including degradation, biocompatibility, and microstructure, and how they translate to inductive microenvironments that stimulate cell infiltration and vascularization to enhance chronic wound healing. As healthcare moves toward precision medicine-based strategies, the potential and therapeutic implications of synthetic, biopolymeric scaffolds as tunable treatment modalities for chronic wounds will be considered. PMID:27547189

  20. Macromolecular recognition: Recognition of polymer side chains by cyclodextrin

    NASA Astrophysics Data System (ADS)

    Hashidzume, Akihito; Harada, Akira

    2015-12-01

    The interaction of cyclodextrins (CD) with water soluble polymers possessing guest residues has been investigated as model systems in biological molecular recognition. The selectivity of interaction of CD with polymer-carrying guest residues is controlled by polymer chains, i.e., the steric effect of polymer main chain, the conformational effect of polymer main chain, and multi-site interaction. Macroscopic assemblies have been also realized based on molecular recognition using polyacrylamide-based gels possessing CD and guest residues.

  1. Multivalent IDP assemblies: Unique properties of LC8-associated, IDP duplex scaffolds.

    PubMed

    Clark, Sarah A; Jespersen, Nathan; Woodward, Clare; Barbar, Elisar

    2015-09-14

    A wide variety of subcellular complexes are composed of one or more intrinsically disordered proteins (IDPs) that are multivalent, flexible, and characterized by dynamic binding of diverse partner proteins. These multivalent IDP assemblies, of broad functional diversity, are classified here into five categories distinguished by the number of IDP chains and the arrangement of partner proteins in the functional complex. Examples of each category are summarized in the context of the exceptional molecular and biological properties of IDPs. One type - IDP duplex scaffolds - is considered in detail. Its unique features include parallel alignment of two IDP chains, formation of new self-associated domains, enhanced affinity for additional bivalent ligands, and ubiquitous binding of the hub protein LC8. For two IDP duplex scaffolds, dynein intermediate chain IC and nucleoporin Nup159, these duplex features, together with the inherent flexibility of IDPs, are central to their assembly and function. A new type of IDP-LC8 interaction, distributed binding of LC8 among multiple IDP recognition sites, is described for Nup159 assembly. PMID:26226419

  2. Logic gates and antisense DNA devices operating on a translator nucleic Acid scaffold.

    PubMed

    Shlyahovsky, Bella; Li, Yang; Lioubashevski, Oleg; Elbaz, Johann; Willner, Itamar

    2009-07-28

    A series of logic gates, "AND", "OR", and "XOR", are designed using a DNA scaffold that includes four "footholds" on which the logic operations are activated. Two of the footholds represent input-recognition strands, and these are blocked by complementary nucleic acids, whereas the other two footholds are blocked by nucleic acids that include the horseradish peroxidase (HRP)-mimicking DNAzyme sequence. The logic gates are activated by either nucleic acid inputs that hybridize to the respective "footholds", or by low-molecular-weight inputs (adenosine monophosphate or cocaine) that yield the respective aptamer-substrate complexes. This results in the respective translocation of the blocking nucleic acids to the footholds carrying the HRP-mimicking DNAzyme sequence, and the concomitant release of the respective DNAzyme. The released product-strands then self-assemble into the hemin/G-quadruplex-HRP-mimicking DNAzyme that biocatalyzes the formation of a colored product and provides an output signal for the different logic gates. The principle of the logic operation is, then, implemented as a possible paradigm for future nanomedicine. The nucleic acid inputs that bind to the blocked footholds result in the translocation of the blocking nucleic acids to the respective footholds carrying the antithrombin aptamer. The released aptamer inhibits, then, the hydrolytic activity of thrombin. The system demonstrates the regulation of a biocatalytic reaction by a translator system activated on a DNA scaffold. PMID:19507821

  3. One-pot synthesis of carbon dots-embedded molecularly imprinted polymer for specific recognition of sterigmatocystin in grains.

    PubMed

    Xu, Longhua; Fang, Guozhen; Pan, Mingfei; Wang, Xuefeng; Wang, Shuo

    2016-03-15

    A novel sensitive fluorescent sensor for determination of sterigmatocystin (ST), which was based on carbon dots-embedded molecularly imprinted polymer (CDs@MIP), was prepared by an efficient one-pot reaction. First, highly blue luminescent CDs were synthesized via a one-step reaction. Then, through a non-hydrolytic sol-gel process, MIP was formed on the CDs surface in the presence of 1,8-dihydroxyanthraquinone as an alternative template molecule to obtain CDs@MIP. The CDs acted as antennas for signal amplification and optical readout, and the MIP coated on the CDs surface provided specific binding sites for ST. The performance of CDs@MIP was compared with that of CDs embedded in non-imprinted polymer (CDs@NIP). CDs@MIP exhibited high selectivity and sensitivity toward ST. Under optimized conditions, the relative fluorescence intensity of CDs@MIP decreased linearly with the concentration of ST from 0.05 to 2.0 mgL(-1) with a detection limit of 0.019 mgL(-1) (S/N=3) and the precision for five replicate detections of 0.10 mgL(-1) ST was 2.31%. The sensor was also used to determine the content of ST in grains with satisfactory results. PMID:26544869

  4. Molecular Phylogenetic and Morphological Evidence Supports Recognition of Gereaua, a New Endemic Genus of Sapindaceae from Madagascar

    PubMed Central

    Buerki, Sven; Lowry, Porter P.; Phillipson, Peter B.; Callmander, Martin W.

    2011-01-01

    A recent worldwide phylogeny of Sapindaceae inferred from nuclear and plastid DNA regions segregated the Malagasy Haplocoelum perrieri Capuron from the African Haplocoelum foliosum (Hiern) Bullock. Additional phylogenetic analyses conducted here (including material of Haplocoelum inopleum Radlk., the generic type) supported the result from the previous analysis and showed that maintaining a broad circumscription of Haplocoelum to include the Malagasy species would render the genus polyphyletic. To maintain monophyly, it is necessary to exclude H. perrieri, which we transfer to a new, monotypic genus, described here as Gereaua. This taxon is easily distinguished from the species retained in Haplocoelum by the following morphological characters: (1) sexually dimorphic inflorescences in racemules (vs. monomorphic inflorescences in fascicule of cymes); (2) 2-locular ovary (vs. 3-locular ovary); (3) rudimentary pistillode in staminate flowers (vs. no pistillode in staminate flowers); (4) corolla with 4 or 5 petals (vs. apetalous); (5) pubescent fruit (vs. glabrous fruit). Relationships between the new genus and its most closely related genera, included in the Macphersonia group, are discussed in light of molecular, morphological and biogeographic evidence. A preliminary threat assessment of Gereaua perrieri using the IUCN Red List criteria indicates a status of Least Concern. PMID:21857766

  5. Molecular and phenotypic data support the recognition of the Wakatobi Flowerpecker (Dicaeum kuehni) from the unique and understudied Sulawesi region.

    PubMed

    Kelly, Seán B A; Kelly, David J; Cooper, Natalie; Bahrun, Andi; Analuddin, Kangkuso; Marples, Nicola M

    2014-01-01

    Accurate estimates of species richness are essential to macroecological and macroevolutionary research, as well as to the effective management and conservation of biodiversity. The resolution of taxonomic relationships is therefore of vital importance. While molecular methods have revolutionised taxonomy, contemporary species delimitation requires an integrative, multi-disciplinary approach. Despite boasting a remarkably high level of endemism, the avifauna of the Sulawesi region of Indonesia remains poorly studied. Previous studies of avian diversity in Sulawesi have focussed predominantly on phenotypic characteristics, thus potentially overlooking any genetically distinct lineages. Grey-sided Flowerpecker Dicaeum celebicum populations from the Wakatobi archipelago were originally described as a separate species from those on nearby mainland Sulawesi. However, for reasons that remain unknown, the Wakatobi populations were reclassified as a subspecies of the mainland form. Combining estimates of genetic divergence with phylogenetic and morphological analyses, we reassessed the status of Wakatobi populations. Our results describe the Wakatobi populations as a separate species to those on mainland Sulawesi; reproductively isolated, genetically and morphologically distinct. We therefore recommend the reclassification of these populations to their original status of Dicaeum kuehni and propose the vernacular name 'Wakatobi Flowerpecker'. In consideration of our findings and the lack of integrative ornithological research within the Sulawesi region, we believe species richness and avian endemism within the region are underestimated. PMID:24896822

  6. Development of small molecule non-peptide formyl peptide receptor (FPR) ligands and molecular modeling of their recognition.

    PubMed

    Schepetkin, I A; Khlebnikov, A I; Giovannoni, M P; Kirpotina, L N; Cilibrizzi, A; Quinn, M T

    2014-01-01

    Formyl peptide receptors (FPRs) are G protein-coupled receptors (GPCRs) expressed on a variety of cell types. These receptors play an important role in the regulation of inflammatory reactions and sensing cellular damage. They have also been implicated in the pathogenesis of various diseases, including neurodegenerative diseases, cataract formation, and atherogenesis. Thus, FPR ligands, both agonists and antagonists, may represent novel therapeutics for modulating host defense and innate immunity. A variety of molecules have been identified as receptor subtype-selective and mixed FPR agonists with potential therapeutic value during last decade. This review describes our efforts along with recent advances in the identification, optimization, biological evaluation, and structure-activity relationship (SAR) analysis of small molecule non-peptide FPR agonists and antagonists, including chiral molecules. Questions regarding the interaction at the molecular level of benzimidazoles, pyrazolones, pyridazin-3(2H)-ones, N-phenylureas and other derivatives with FPR1 and FPR2 are discussed. Application of computational models for virtual screening and design of FPR ligands is also considered. PMID:24350845

  7. Molecular recognition mechanism of peptide chain bound to the tRNA(Lys3) anticodon loop in silico.

    PubMed

    Xiao, Xingqing; Agris, Paul F; Hall, Carol K

    2015-01-01

    The mechanism by which proteins recognize and bind the post-transcriptional modifications of RNAs is unknown, yet these interactions play important functions in biology. Atomistic molecular dynamics simulations were performed to examine the folding of the model peptide chain -RVTHHAFLGAHRTVG- and the complex formed by the folded peptide with the native anticodon stem and loop of the human tRNA(Lys3) (hASL(Lys3)) in order to explore the binding mechanism. By analyzing and comparing two folded conformations of this peptide obtained from the folding simulation, we found that the van der Waals (VDW) energy is necessary for the thermal stability of the peptide, and the charge-charge (ELE + EGB) energy is crucial for determining the three-dimensional folded structure of the peptide backbone. Subsequently, two conformations of the peptide were employed to investigate their binding behaviors to hASL(Lys3). The metastable folded peptide was found to bind to hASL(Lys3) much easier than the stable folded peptide in the binding simulations. An energetic analysis reveals that the VDW energy favors the binding, whereas the ELE + EGB energies disfavor the binding. Arginines on the peptide preferentially attract the phosphate backbone via the inter-chain ELE + EGB interaction, significantly contributing to the binding affinity. The hydrophobic phenylalanine interacts with the anticodon loop of hASL(Lys3) via the inter-chain VDW interaction, significantly contributing to the binding specificity. PMID:24417415

  8. Differentiation of peptide molecular recognition by phospholipase C gamma-1 Src homology-2 domain and a mutant Tyr phosphatase PTP1bC215S.

    PubMed Central

    MacLean, D.; Sefler, A. M.; Zhu, G.; Decker, S. J.; Saltiel, A. R.; Singh, J.; McNamara, D.; Dobrusin, E. M.; Sawyer, T. K.

    1995-01-01

    Activated epidermal growth factor receptor (EGFR) undergoes autophosphorylation on several cytoplasmic tyrosine residues, which may then associate with the src homology-2 (SH2) domains of effector proteins such as phospholipase C gamma-1 (PLC gamma-1). Specific phosphotyrosine (pTyr)-modified EGFR fragment peptides can inhibit this intermolecular binding between activated EGFR and a tandem amino- and carboxy-terminal (N/C) SH2 protein construct derived from PLC gamma-1. In this study, we further explored the molecular recognition of phosphorylated EGFR988-998 (Asp-Ala-Asp-Glu-pTyr-Leu-Ile-Pro-Gln-Gln-Gly, I) by PLC gamma-1 N/C SH2 in terms of singular Ala substitutions for amino acid residues N- and C-terminal to the pTyr (P site) of phosphopeptide I. Comparison of the extent to which these phosphopeptides inhibited binding of PLC gamma-1 N/C SH2 to activated EGFR showed the critical importance of amino acid side chains at positions P+2 (Ile994), P+3 (Pro995), and P+4 (Gln996). Relative to phosphopeptide I, multiple Ala substitution throughout the N-terminal sequence, N-terminal sequence, N-terminal truncation, or dephosphorylation of pTyr each resulted in significantly decreased binding to PLC gamma-1 N/C SH2. These structure-activity results were analyzed by molecular modeling studies of the predicted binding of phosphopeptide I to each the N- and C-terminal SH2 domains of PLC gamma-1.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:7773170

  9. Mathematical Abstraction through Scaffolding

    ERIC Educational Resources Information Center

    Ozmantar, Mehmet Fatih; Roper, Tom

    2004-01-01

    This paper examines the role of scaffolding in the process of abstraction. An activity-theoretic approach to abstraction in context is taken. This examination is carried out with reference to verbal protocols of two 17 year-old students working together on a task connected to sketching the graph of |f|x|)|. Examination of the data suggests that…

  10. Molecular basis of lipo-chitooligosaccharide recognition by the lysin motif receptor-like kinase LYR3 in legumes.

    PubMed

    Malkov, Nikita; Fliegmann, Judith; Rosenberg, Charles; Gasciolli, Virginie; Timmers, Antonius C J; Nurisso, Alessandra; Cullimore, Julie; Bono, Jean-Jacques

    2016-05-15

    LYR3 [LysM (lysin motif) receptor-like kinase 3] of Medicago truncatula is a high-affinity binding protein for symbiotic LCO (lipo-chitooligosaccharide) signals, produced by rhizobia bacteria and arbuscular mycorrhizal fungi. The present study shows that LYR3 from several other legumes, but not from two Lupinus species which are incapable of forming the mycorrhizal symbiosis, bind LCOs with high affinity and discriminate them from COs (chitooligosaccharides). The biodiversity of these proteins and the lack of binding to the Lupinus proteins were used to identify features required for high-affinity LCO binding. Swapping experiments between each of the three LysMs of the extracellular domain of the M. truncatula and Lupinus angustifolius LYR3 proteins revealed the crucial role of the third LysM in LCO binding. Site-directed mutagenesis identified a tyrosine residue, highly conserved in all LYR3 LCO-binding proteins, which is essential for high-affinity binding. Molecular modelling suggests that it may be part of a hydrophobic tunnel able to accommodate the LCO acyl chain. The lack of conservation of these features in the binding site of plant LysM proteins binding COs provides a mechanistic explanation of how LCO recognition might differ from CO perception by structurally related LysM receptors. PMID:26987814

  11. Facile synthesis of mono-6-amino-6-deoxy-alpha-, beta-, gamma-cyclodextrin hydrochlorides for molecular recognition, chiral separation and drug delivery.

    PubMed

    Tang, Weihua; Ng, Siu-Choon

    2008-01-01

    We describe a protocol for the synthesis of mono-6-amino-6-deoxy-cyclodextrin hydrochloride (CD-NH3Cl), applicable to alpha-, beta- and gamma-cyclodextrin. These structurally simplest, highly water-soluble cationic cyclodextrins can be widely used in molecular recognition, chiral separation and drug delivery studies. Starting from commercially available chemicals, CD-NH3Cl is synthesized in four steps: (i) selective tosylation of cyclodextrin by the use of p-toluenesulfonyl chloride to afford mono-6-(p-toluenesulfonyl)-6-deoxy-cyclodextrin (Ts-CD); (ii) azide substitution of Ts-CD with sodium azide to afford mono-6-azido-6-deoxy-cyclodextrin (CD-N3); (iii) reduction of CD-N3 with triphenylphospine followed by hydrolysis to prepare mono-6-amino-6-deoxy-cyclodextrin (CD-NH2); and (iv) treatment of CD-NH2 with hydrochloric acid to afford the titled CD-NH3Cl with good yield. The overall protocol requires approximately 2 weeks. PMID:18388952

  12. In Vitro Selection of a Single-Stranded DNA Molecular Recognition Element against Clostridium difficile Toxin B and Sensitive Detection in Human Fecal Matter

    PubMed Central

    Maher, Eamonn; Williams, Ryan M.; Sooter, Letha J.

    2015-01-01

    Toxin B is one of the major virulence factors of Clostridium difficile, a bacterium that is responsible for a significant number of diarrhea cases in acute care settings. Due to the prevalence of C. difficile induced diarrhea, rapid and correct diagnosis is crucial in the disease management. In this study, we have employed a stringent in vitro selection method to identify single-stranded DNA molecular recognition elements (MRE) specific for toxin B. At the end of the 12-round selection, one MRE with high affinity (Kd = 47.3 nM) for toxin B was identified. The selected MRE demonstrated low cross binding activities on negative targets: bovine serum albumin, Staphylococcus aureus alpha toxin, Pseudomonas aeruginosa exotoxin A, and cholera toxin of Vibrio cholera. A modified sandwich ELISA assay was developed utilizing the selected ssDNA MRE as the antigen capturing element and achieved a sensitive detection of 50 nM of toxin B in human fecal preparations. PMID:25734010

  13. In Vitro Selection of Single-Stranded DNA Molecular Recognition Elements against S. aureus Alpha Toxin and Sensitive Detection in Human Serum

    PubMed Central

    Hong, Ka L.; Battistella, Luisa; Salva, Alysia D.; Williams, Ryan M.; Sooter, Letha J.

    2015-01-01

    Alpha toxin is one of the major virulence factors secreted by Staphylococcus aureus, a bacterium that is responsible for a wide variety of infections in both community and hospital settings. Due to the prevalence of S. aureus related infections and the emergence of methicillin-resistant S. aureus, rapid and accurate diagnosis of S. aureus infections is crucial in benefiting patient health outcomes. In this study, a rigorous Systematic Evolution of Ligands by Exponential Enrichment (SELEX) variant previously developed by our laboratory was utilized to select a single-stranded DNA molecular recognition element (MRE) targeting alpha toxin with high affinity and specificity. At the end of the 12-round selection, the selected MRE had an equilibrium dissociation constant (Kd) of 93.7 ± 7.0 nM. Additionally, a modified sandwich enzyme-linked immunosorbent assay (ELISA) was developed by using the selected ssDNA MRE as the toxin-capturing element and a sensitive detection of 200 nM alpha toxin in undiluted human serum samples was achieved. PMID:25633102

  14. Selection of Single-Stranded DNA Molecular Recognition Elements against Exotoxin A Using a Novel Decoy-SELEX Method and Sensitive Detection of Exotoxin A in Human Serum

    PubMed Central

    Hong, Ka Lok; Yancey, Kailey; Battistella, Luisa; Williams, Ryan M.; Hickey, Katherine M.; Bostick, Chris D.; Gannett, Peter M.; Sooter, Letha J.

    2015-01-01

    Exotoxin A is one of the virulence factors of Pseudomonas aeruginosa, a bacterium that can cause infections resulting in adverse health outcomes and increased burden to health care systems. Current methods of diagnosing P. aeruginosa infections are time consuming and can require significant preparation of patient samples. This study utilized a novel variation of the Systematic Evolution of Ligand by Exponential Enrichment, Decoy-SELEX, to identify an Exotoxin A specific single-stranded DNA (ssDNA) molecular recognition element (MRE). Its emphasis is on increasing stringency in directing binding toward free target of interest and at the same time decreasing binding toward negative targets. A ssDNA MRE with specificity and affinity was identified after fourteen rounds of Decoy-SELEX. Utilizing surface plasmon resonance measurements, the determined equilibrium dissociation constant (Kd) of the MRE is between 4.2 µM and 4.5 µM, and is highly selective for Exotoxin A over negative targets. A ssDNA MRE modified sandwich enzyme-linked immunosorbent assay (ELISA) has been developed and achieved sensitive detection of Exotoxin A at nanomolar concentrations in human serum. This study has demonstrated the proof-of-principle of using a ssDNA MRE as a clinical diagnostic tool. PMID:26636098

  15. Molecular cloning and characterisation of a pattern recognition molecule, lipopolysaccharide- and beta-1,3-glucan binding protein (LGBP) from the white shrimp Litopenaeus vannamei.

    PubMed

    Cheng, Winton; Liu, Chun-Hung; Tsai, Chiung-Hui; Chen, Jiann-Chu

    2005-04-01

    A lipopolysaccharide- and beta-1,3-glucan binding protein (LGBP) cDNA was cloned from the haemocyte and hepatopancreas of white shrimp Litopenaeus vannamei using oligonucleotide primers and RT-PCR. Both 3'- and 5'-regions were isolated by rapid amplification of cDNA end RACE method. Analysis of nucleotide sequence revealed that the cDNA clone has an open reading frame of 1101 bp encoding a protein of 367 amino acids including a 17 amino acid signal peptide. The calculated molecular mass of the mature proteins (350 amino acids) is 39.92 kDa with an estimated pI of 4.37. Two putative integrin binding motifs (cell adhesion site), RGD (Arg-Gly-Asp) and a potential recognition motif for beta- (1-->3) linkage of polysaccharides were observed in the LGBP. Sequence comparison showed that LGBP deduced amino acid of L. vannamei has an overall similarity of 95%, 92% and 61% to that of blue shrimp Litopenaeus stylirostris LGBP, tiger shrimp Penaeus monodon BGBP and crayfish Pacifastacus leniusculus LGBP, respectively. Quantitative real-time RT-PCR analysis showed that LGBP transcript in haemocyte of L. vannamei increased in 3- and 6-h post Vibrio alginolyticus injection. PMID:15561560

  16. Crossreactive recognition of viral, self, and bacterial peptide ligands by human class I-restricted cytotoxic T lymphocyte clonotypes: Implications for molecular mimicry in autoimmune disease

    PubMed Central

    Misko, Ihor S.; Cross, Simone M.; Khanna, Rajiv; Elliott, Suzanne L.; Schmidt, Christopher; Pye, Stephanie J.; Silins, Sharon L.

    1999-01-01

    The immunodominant, CD8+ cytotoxic T lymphocyte (CTL) response to the HLA-B8-restricted peptide, RAKFKQLL, located in the Epstein–Barr virus immediate-early antigen, BZLF1, is characterized by a diverse T cell receptor (TCR) repertoire. Here, we show that this diversity can be partitioned on the basis of crossreactive cytotoxicity patterns involving the recognition of a self peptide—RSKFRQIV—located in a serine/threonine kinase and a bacterial peptide—RRKYKQII—located in Staphylococcus aureus replication initiation protein. Thus CTL clones that recognized the viral, self, and bacterial peptides expressed a highly restricted αβ TCR phenotype. The CTL clones that recognized viral and self peptides were more oligoclonal, whereas clones that strictly recognized the viral peptide displayed a diverse TCR profile. Interestingly, the self and bacterial peptides equally were substantially less effective than the cognate viral peptide in sensitizing target cell lysis, and also resulted only in a weak reactivation of memory CTLs in limiting dilution assays, whereas the cognate peptide was highly immunogenic. The described crossreactions show that human antiviral, CD8+ CTL responses can be shaped by peptide ligands derived from autoantigens and environmental bacterial antigens, thereby providing a firm structural basis for molecular mimicry involving class I-restricted CTLs in the pathogenesis of autoimmune disease. PMID:10051632

  17. Molecular Basis of Histone Tail Recognition by Human TIP5 PHD Finger and Bromodomain of the Chromatin Remodeling Complex NoRC

    PubMed Central

    Tallant, Cynthia; Valentini, Erica; Fedorov, Oleg; Overvoorde, Lois; Ferguson, Fleur M.; Filippakopoulos, Panagis; Svergun, Dmitri I.; Knapp, Stefan; Ciulli, Alessio

    2015-01-01

    Summary Binding of the chromatin remodeling complex NoRC to RNA complementary to the rDNA promoter mediates transcriptional repression. TIP5, the largest subunit of NoRC, is involved in recruitment to rDNA by interactions with promoter-bound TTF-I, pRNA, and acetylation of H4K16. TIP5 domains that recognize posttranslational modifications on histones are essential for recruitment of NoRC to chromatin, but how these reader modules recognize site-specific histone tails has remained elusive. Here, we report crystal structures of PHD zinc finger and bromodomains from human TIP5 and BAZ2B in free form and bound to H3 and/or H4 histones. PHD finger functions as an independent structural module in recognizing unmodified H3 histone tails, and the bromodomain prefers H3 and H4 acetylation marks followed by a key basic residue, KacXXR. Further low-resolution analyses of PHD-bromodomain modules provide molecular insights into their trans histone tail recognition, required for nucleosome recruitment and transcriptional repression of the NoRC complex. PMID:25533489

  18. Deciphering the Non-Equivalence of Serine and Threonine O-Glycosylation Points: Implications for Molecular Recognition of the Tn Antigen by an anti-MUC1 Antibody**

    PubMed Central

    Martínez-Sáez, Nuria; Castro-López, Jorge; Valero-González, Jessika; Madariaga, David; Compañón, Ismael; Somovilla, Víctor J; Salvadó, Míriam; Asensio, Juan L; Jiménez-Barbero, Jesús; Avenoza, Alberto; Busto, Jesús H; Bernardes, Gonçalo J L; Peregrina, Jesús M; Hurtado-Guerrero, Ramón; Corzana, Francisco

    2015-01-01

    The structural features of MUC1-like glycopeptides bearing the Tn antigen (α-O-GalNAc-Ser/Thr) in complex with an anti MUC-1 antibody are reported at atomic resolution. For the α-O-GalNAc-Ser derivative, the glycosidic linkage adopts a high-energy conformation, barely populated in the free state. This unusual structure (also observed in an α-S-GalNAc-Cys mimic) is stabilized by hydrogen bonds between the peptidic fragment and the sugar. The selection of a particular peptide structure by the antibody is thus propagated to the carbohydrate through carbohydrate/peptide contacts, which force a change in the orientation of the sugar moiety. This seems to be unfeasible in the α-O-GalNAc-Thr glycopeptide owing to the more limited flexibility of the side chain imposed by the methyl group. Our data demonstrate the non-equivalence of Ser and Thr O-glycosylation points in molecular recognition processes. These features provide insight into the occurrence in nature of the APDTRP epitope for anti-MUC1 antibodies. PMID:26118689

  19. Molecular basis of histone tail recognition by human TIP5 PHD finger and bromodomain of the chromatin remodeling complex NoRC.

    PubMed

    Tallant, Cynthia; Valentini, Erica; Fedorov, Oleg; Overvoorde, Lois; Ferguson, Fleur M; Filippakopoulos, Panagis; Svergun, Dmitri I; Knapp, Stefan; Ciulli, Alessio

    2015-01-01

    Binding of the chromatin remodeling complex NoRC to RNA complementary to the rDNA promoter mediates transcriptional repression. TIP5, the largest subunit of NoRC, is involved in recruitment to rDNA by interactions with promoter-bound TTF-I, pRNA, and acetylation of H4K16. TIP5 domains that recognize posttranslational modifications on histones are essential for recruitment of NoRC to chromatin, but how these reader modules recognize site-specific histone tails has remained elusive. Here, we report crystal structures of PHD zinc finger and bromodomains from human TIP5 and BAZ2B in free form and bound to H3 and/or H4 histones. PHD finger functions as an independent structural module in recognizing unmodified H3 histone tails, and the bromodomain prefers H3 and H4 acetylation marks followed by a key basic residue, KacXXR. Further low-resolution analyses of PHD-bromodomain modules provide molecular insights into their trans histone tail recognition, required for nucleosome recruitment and transcriptional repression of the NoRC complex. PMID:25533489

  20. Co-conformational Exchange Triggered by Molecular Recognition in a Di(acylamino)pyridine-Based Molecular Shuttle Containing Two Pyridine Rings at the Macrocycle.

    PubMed

    Martinez-Cuezva, Alberto; Carro-Guillen, Fernando; Pastor, Aurelia; Marin-Luna, Marta; Orenes, Raul-Angel; Alajarin, Mateo; Berna, Jose

    2016-06-17

    We describe the incorporation of endo-pyridine units into the tetralactam ring of di(acylamino)pyridine-based rotaxanes. This macrocycle strongly associates with the linear interlocked component as confirmed by X-ray diffraction studies of rotaxane 2 b. Dynamic NMR studies of 2 b in solution revealed a rotational energy barrier that was higher than that of the related rotaxane 2 a, which lacks of pyridine rings in the macrocycle. The macrocycle distribution of the molecular shuttle 4 b, containing two endo-pyridine rings, shows that the major co-conformer is that with the cyclic component sitting over the di(acylamino)pyridine station. DFT calculations also support the marked preference of the ring for occupying the heterocyclic binding site. The association of N-hexylthymine with the di(acylamino)pyridine binding site of 4 b led to the formation of a rare 'S'-shaped co-conformer in which the tetralactam ring interacts simultaneously with both stations of the thread. PMID:26698891

  1. Advances in skin regeneration: application of electrospun scaffolds.

    PubMed

    Norouzi, Mohammad; Boroujeni, Samaneh Moghadasi; Omidvarkordshouli, Noushin; Soleimani, Masoud

    2015-06-01

    The paucity of cellular and molecular signals essential for normal wound healing makes severe dermatological ulcers stubborn to heal. The novel strategies of skin regenerative treatments are focused on the development of biologically responsive scaffolds accompanied by cells and multiple biomolecules resembling structural and biochemical cues of the natural extracellular matrix (ECM). Electrospun nanofibrous scaffolds provide similar architecture to the ECM leading to enhancement of cell adhesion, proliferation, migration and neo tissue formation. This Review surveys the application of biocompatible natural, synthetic and composite polymers to fabricate electrospun scaffolds as skin substitutes and wound dressings. Furthermore, the application of biomolecules and therapeutic agents in the nanofibrous scaffolds viz growth factors, genes, antibiotics, silver nanoparticles, and natural medicines with the aim of ameliorating cellular behavior, wound healing, and skin regeneration are discussed. PMID:25721694

  2. Synthesis and characterization of liquid-crystalline supramolecular architecture by a combination of molecular recognition and polymerization reaction

    NASA Astrophysics Data System (ADS)

    Ahn, Cheol-Hee

    for polymers with controlled cylindrical and spherical shapes which assemble into thermotropic liquid crystalline phase. The molecular design, synthesis, characterization and polymerization of these bulky monomers and polymers requires a combination of supramolecular chemistry, liquid crystallinity and polymer science. This thesis will provide an entry into the design of visualizable polymeric materials with dramatically different properties which are determined by their shape. Cylindrical and spherical polymers obtained from flexible backbones jacketed with dendritic coats will access a mechanism to control the conformation and rigidity of their flexible chains. The rigidity of cylindrical polymers can be manipulated via their diameter, internal structure and conformational restrictions. Mechanisms to change in a reversible and controlled way to change the shape of the polymer from rod-like to spherical is elaborated and this work can open a new technological concepts in areas such as fibers, films, membranes, electronic, photonic and controlled release systems.

  3. The development and implantation of a biologically derived allograft scaffold.

    PubMed

    Nowocin, Anna K; Southgate, Aaron; Shurey, Sandra; Sibbons, Paul; Gabe, Simon M; Ansari, Tahera

    2016-02-01

    Biologically derived scaffolds are becoming viable treatment options for tissue/organ repair and regeneration. A continuing hurdle is the need for a functional blood supply to and from the implanted scaffold. We have addressed this problem by constructing an acellular ileal scaffold with an attached vascular network suitable for implantation and immediate reperfusion with the host's blood. Using a vascular perfusion approach, a segment of porcine ileum up to 30 cm long, together with its attached vasculature, was decellularized as a single entity. The quality of the decellularized scaffold was assessed histologically and using molecular tools. To establish vascular perfusion potentials of the scaffold, a right-sided nephrectomy and end-to-end anastomosis of the decellularized scaffold's vasculature to a renal artery and vein were performed in a pig of similar size to the donor animal. Lengths of ileal scaffold, together with its attached vasculature, were successfully decellularized, with no evidence of intact cells/nuclear material or collagen degradation. The scaffold's decellularized vascular network demonstrated optimum perfusion at 1, 2 and 24 h post-implantation and the mesenteric arcade remained patent throughout the assessment. The 1, 2 and 24 h explanted scaffolds demonstrated signs of cellular attachment, with cells positive for CD68 and CD133 on the vascular luminal aspect. It is possible to decellularize clinically relevant lengths of small intestine, together with the associated vasculature, as a single segment. The functional vascular network may represent a route for recellularization for future regeneration of bowel tissue for patients with short bowel syndrome. Copyright © 2013 John Wiley & Sons, Ltd. PMID:23554406

  4. Facilitated receptor-recognition and enhanced bioactivity of bone morphogenetic protein-2 on magnesium-substituted hydroxyapatite surface

    NASA Astrophysics Data System (ADS)

    Huang, Baolin; Yuan, Yuan; Li, Tong; Ding, Sai; Zhang, Wenjing; Gu, Yuantong; Liu, Changsheng

    2016-04-01

    Biomaterial surface functionalized with bone morphogenetic protein-2 (BMP-2) is a promising approach to fabricating successful orthopedic implants/scaffolds. However, the bioactivity of BMP-2 on material surfaces is still far from satisfactory and the mechanism of related protein-surface interaction remains elusive. Based on the most widely used bone-implants/scaffolds material, hydroxyapatite (HAP), we developed a matrix of magnesium-substituted HAP (Mg-HAP, 2.2 at% substitution) to address these issues. Further, we investigated the adsorption dynamics, BMPRs-recruitment, and bioactivity of recombinant human BMP-2 (rhBMP-2) on the HAP and Mg-HAP surfaces. To elucidate the mechanism, molecular dynamic simulations were performed to calculate the preferred orientations, conformation changes, and cysteine-knot stabilities of adsorbed BMP-2 molecules. The results showed that rhBMP-2 on the Mg-HAP surface exhibited greater bioactivity, evidenced by more facilitated BMPRs-recognition and higher ALP activity than on the HAP surface. Moreover, molecular simulations indicated that BMP-2 favoured distinct side-on orientations on the HAP and Mg-HAP surfaces. Intriguingly, BMP-2 on the Mg-HAP surface largely preserved the active protein structure evidenced by more stable cysteine-knots than on the HAP surface. These findings explicitly clarify the mechanism of BMP-2-HAP/Mg-HAP interactions and highlight the promising application of Mg-HAP/BMP-2 matrixes in bone regeneration implants/scaffolds.

  5. Facilitated receptor-recognition and enhanced bioactivity of bone morphogenetic protein-2 on magnesium-substituted hydroxyapatite surface

    PubMed Central

    Huang, Baolin; Yuan, Yuan; Li, Tong; Ding, Sai; Zhang, Wenjing; Gu, Yuantong; Liu, Changsheng

    2016-01-01

    Biomaterial surface functionalized with bone morphogenetic protein-2 (BMP-2) is a promising approach to fabricating successful orthopedic implants/scaffolds. However, the bioactivity of BMP-2 on material surfaces is still far from satisfactory and the mechanism of related protein-surface interaction remains elusive. Based on the most widely used bone-implants/scaffolds material, hydroxyapatite (HAP), we developed a matrix of magnesium-substituted HAP (Mg-HAP, 2.2 at% substitution) to address these issues. Further, we investigated the adsorption dynamics, BMPRs-recruitment, and bioactivity of recombinant human BMP-2 (rhBMP-2) on the HAP and Mg-HAP surfaces. To elucidate the mechanism, molecular dynamic simulations were performed to calculate the preferred orientations, conformation changes, and cysteine-knot stabilities of adsorbed BMP-2 molecules. The results showed that rhBMP-2 on the Mg-HAP surface exhibited greater bioactivity, evidenced by more facilitated BMPRs-recognition and higher ALP activity than on the HAP surface. Moreover, molecular simulations indicated that BMP-2 favoured distinct side-on orientations on the HAP and Mg-HAP surfaces. Intriguingly, BMP-2 on the Mg-HAP surface largely preserved the active protein structure evidenced by more stable cysteine-knots than on the HAP surface. These findings explicitly clarify the mechanism of BMP-2-HAP/Mg-HAP interactions and highlight the promising application of Mg-HAP/BMP-2 matrixes in bone regeneration implants/scaffolds. PMID:27075233

  6. Bilayer porous scaffold based on poly-(ɛ-caprolactone) nanofibrous membrane and gelatin sponge for favoring cell proliferation

    NASA Astrophysics Data System (ADS)

    Zhou, Zhihua; Zhou, Yang; Chen, Yiwang; Nie, Huarong; Wang, Yang; Li, Fan; Zheng, Yan

    2011-12-01

    Electrospun poly-(ɛ-caprolactone) (PCL) nanofibers has been widely used in the medical prosthesis. However, poor hydrophilicity and the lack of natural recognition sites for covalent cell-recognition signal molecules to promote cell attachment have limited its utility as tissue scaffolds. In this study, Bilayer porous scaffolds based on PCL electrospun membranes and gelatin (GE) sponges were fabricated through soft hydrolysis of PCL electrospun followed by grafting gelatin onto the fiber surface, through crosslinking and freeze drying treatment of additional gelatin coat and grafted gelatin surface. GE sponges were stably anchored on PCL membrane surface with the aid of grafted GE molecules. The morphologies of bilayer porous scaffolds were observed through SEM. The contact angle of the scaffolds was 0°, the mechanical properties of scaffolds were measured by tensile test, Young's moduli of PCL scaffolds before and after hydrolysis are 66-77.3 MPa and 62.3-75.4 MPa, respectively. Thus, the bilayer porous scaffolds showed excellent hydrophilic surface and desirable mechanical strength due to the soft hydrolysis and GE coat. The cell culture results showed that the adipose derived mesenchymal stem cells did more favor to adhere and grow on the bilayer porous scaffolds than on PCL electrospun membranes. The better cell affinity of the final bilayer scaffolds not only attributed to the surface chemistry but also the introduction of bilayer porous structure.

  7. Protein Scaffolding for Small Molecule Catalysts

    SciTech Connect

    Baker, David

    2014-09-14

    We aim to design hybrid catalysts for energy production and storage that combine the high specificity, affinity, and tunability of proteins with the potent chemical reactivities of small organometallic molecules. The widely used Rosetta and RosettaDesign methodologies will be extended to model novel protein / small molecule catalysts in which one or many small molecule active centers are supported and coordinated by protein scaffolding. The promise of such hybrid molecular systems will be demonstrated with the nickel-phosphine hydrogenase of DuBois et. al.We will enhance the hydrogenase activity of the catalyst by designing protein scaffolds that incorporate proton relays and systematically modulate the local environment of the catalyticcenter. In collaboration with DuBois and Shaw, the designs will be experimentally synthesized and characterized.

  8. Evolution Stings: The Origin and Diversification of Scorpion Toxin Peptide Scaffolds

    PubMed Central

    Sunagar, Kartik; Undheim, Eivind A. B.; Chan, Angelo H. C.; Koludarov, Ivan; Muñoz-Gómez, Sergio A.; Antunes, Agostinho; Fry, Bryan G.

    2013-01-01

    The episodic nature of natural selection and the accumulation of extreme sequence divergence in venom-encoding genes over long periods of evolutionary time can obscure the signature of positive Darwinian selection. Recognition of the true biocomplexity is further hampered by the limited taxon selection, with easy to obtain or medically important species typically being the subject of intense venom research, relative to the actual taxonomical diversity in nature. This holds true for scorpions, which are one of the most ancient terrestrial venomous animal lineages. The family Buthidae that includes all the medically significant species has been intensely investigated around the globe, while almost completely ignoring the remaining non-buthid families. Australian scorpion lineages, for instance, have been completely neglected, with only a single scorpion species (Urodacus yaschenkoi) having its venom transcriptome sequenced. Hence, the lack of venom composition and toxin sequence information from an entire continent’s worth of scorpions has impeded our understanding of the molecular evolution of scorpion venom. The molecular origin, phylogenetic relationships and evolutionary histories of most scorpion toxin scaffolds remain enigmatic. In this study, we have sequenced venom gland transcriptomes of a wide taxonomical diversity of scorpions from Australia, including buthid and non-buthid representatives. Using state-of-art molecular evolutionary analyses, we show that a majority of CSα/β toxin scaffolds have experienced episodic influence of positive selection, while most non-CSα/β linear toxins evolve under the extreme influence of negative selection. For the first time, we have unraveled the molecular origin of the major scorpion toxin scaffolds, such as scorpion venom single von Willebrand factor C-domain peptides (SV-SVC), inhibitor cystine knot (ICK), disulphide-directed beta-hairpin (DDH), bradykinin potentiating peptides (BPP), linear non-disulphide bridged

  9. Scaffolding Student Participation in Mathematical Practices

    ERIC Educational Resources Information Center

    Moschkovich, Judit N.

    2015-01-01

    The concept of scaffolding can be used to describe various types of adult guidance, in multiple settings, across different time scales. This article clarifies what we mean by scaffolding, considering several questions specifically for scaffolding in mathematics: What theoretical assumptions are framing scaffolding? What is being scaffolded? At…

  10. Bioresorbable Scaffolds for Atheroregression: Understanding of Transient Scaffolding.

    PubMed

    Kharlamov, Alexander N

    2016-01-01

    This review focuses on the clinical and biological features of the bioresorbable scaffolds in interventional cardiology highlighting scientific achievements and challenges of the transient scaffolding with Absorb BVS. Special attention is granted to the vascular biology pathways which, involved in the resorption of scaffold, artery remodeling and mechanisms of Glagovian atheroregression setting the stage for subsequent clinical applications. Twenty five years ago Glagov described the phenomenon of limited external elastic membrane enlargement in response to an increase in plaque burden. We believe this threshold becomes the target for development of strategies that reverse atherosclerosis, and particularly transient scaffolding has a potential to be a tool to ultimately conquer atherosclerosis. PMID:26818488

  11. Biologically inspired growth of hydroxyapatite crystals on bio-organics-defined scaffolds

    SciTech Connect

    Yang, Chunrong; Li, Yuli; Nan, Kaihui

    2013-03-15

    Graphical abstract: Petal-like crystals were observed to form on the surface of the BG/COL/ChS scaffolds. Highlights: ► Porous scaffolds were prepared using bioglass, collagen and chondroitin sulfate. ► Highly oriented HA crystals were grown on scaffolds using simulated body fluids ► The microstructure and orientation of HA were explained by molecular configuration. - Abstract: Several bio-organics-defined composite scaffolds were prepared using 58s-bioglass (BG), collagen (Col) and chondroitin sulfate (ChS). These scaffolds possess highly porous structure. X-ray diffraction of these scaffolds strongly indicated that hydroxyapatite (HA) crystals formed on their surfaces in simulated body fluids within 3 d, and similar formation process of crystals could be obtained on BG/Col and BG/Col/ChS scaffolds. The morphology and structure of the crystals were further examined by scanning electron microscopy. The results obtained indicate that an apatite with petal-like structure similar to that found on BG/Col scaffolds can be produced on BG/Col/ChS scaffolds through biomimetic synthesis, while that on BG/ChS scaffolds took place differently. The differences could be explained by self-assembly processes and the different macromolecular configurations of the Col and ChS fibrils which self-assemble spontaneously into their fibers. On the other hand, the bio-organics-defined composites have good cell biocompability. The results may be applicable to develop tailored biomaterials for peculiar bone substitute.

  12. Molecular dynamics simulations elucidate the mode of protein recognition by Skp1 and the F-box domain in the SCF complex.

    PubMed

    Chandra Dantu, Sarath; Nathubhai Kachariya, Nitin; Kumar, Ashutosh

    2016-01-01

    Polyubiquitination of the target protein by a ubiquitin transferring machinery is key to various cellular processes. E3 ligase Skp1-Cul1-F-box (SCF) is one such complex which plays crucial role in substrate recognition and transfer of the ubiquitin molecule. Previous computational studies have focused on S-phase kinase-associated protein 2 (Skp2), cullin, and RING-finger proteins of this complex, but the roles of the adapter protein Skp1 and F-box domain of Skp2 have not been determined. Using sub-microsecond molecular dynamics simulations of full-length Skp1, unbound Skp2, Skp2-Cks1 (Cks1: Cyclin-dependent kinases regulatory subunit 1), Skp1-Skp2, and Skp1-Skp2-Cks1 complexes, we have elucidated the function of Skp1 and the F-box domain of Skp2. We found that the L16 loop of Skp1, which was deleted in previous X-ray crystallography studies, can offer additional stability to the ternary complex via its interactions with the C-terminal tail of Skp2. Moreover, Skp1 helices H6, H7, and H8 display vivid conformational flexibility when not bound to Skp2, suggesting that these helices can recognize and lock the F-box proteins. Furthermore, we observed that the F-box domain could rotate (5°-129°), and that the binding partner determined the degree of conformational flexibility. Finally, Skp1 and Skp2 were found to execute a domain motion in Skp1-Skp2 and Skp1-Skp2-Cks1 complexes that could decrease the distance between ubiquitination site of the substrate and the ubiquitin molecule by 3 nm. Thus, we propose that both the F-box domain of Skp2 and Skp1-Skp2 domain motions displaying preferential conformational control can together facilitate polyubiquitination of a wide variety of substrates. PMID:26573739

  13. Adsorption characteristics, recognition properties, and preliminary application of nordihydroguaiaretic acid molecularly imprinted polymers prepared by sol-gel surface imprinting technology

    NASA Astrophysics Data System (ADS)

    Liao, Sen; Zhang, Wen; Long, Wei; Hou, Dan; Yang, Xuechun; Tan, Ni

    2016-02-01

    In this paper, a new core-shell composite of nordihydroguaiaretic acid (NDGA) molecularly imprinted polymers layer-coated silica gel (MIP@SiO2) was prepared through sol-gel technique and applied as a material for extraction of NDGA from Ephedra. It was synthesized using NDGA as the template molecule, γ-aminopropyltriethoxysilane (APTS) and methyltriethoxysilane (MTEOS) as the functional monomers, tetraethyl orthosilicate (TEOS) as the cross-linker and ethanol as the porogenic solvent in the surface of silica. The non-imprinted polymers layer-coated silica gel (NIP@SiO2) were prepared with the same procedure, but with the absence of template molecule. In addition, the optimum adsorption affinity occurred when the molar ratio of NDGA:APTS:MTEOS:TEOS was 1:6:2:80. The prepared MIP@SiO2 and NIP@SiO2 were analyzed by scanning electron microscopy (SEM), thermogravimetric analysis (TGA), and Fourier transform-infrared spectroscopy (FT-IR). Their affinity properties to NDGA were evaluated through dynamic adsorption, static adsorption, and selective recognition experiments, and the results showed the saturated adsorption capacity of MIP@SiO2 could reach to 5.90 mg g-1, which was two times more than that of NIP@SiO2. High performance liquid chromatography (HPLC) was used to evaluate the extraction of NDGA from the medicinal plant ephedra by the above prepared materials, and the results indicated that the MIP@SiO2 had potential application in separation of the natural active component NDGA from medicinal plants.

  14. Molecular dynamics investigations of ozone on an ab initio potential energy surface with the utilization of pattern-recognition neural network for accurate determination of product formation.

    PubMed

    Le, Hung M; Dinh, Thach S; Le, Hieu V

    2011-10-13

    The singlet-triplet transformation and molecular dissociation of ozone (O(3)) gas is investigated by performing quasi-classical molecular dynamics (MD) simulations on an ab initio potential energy surface (PES) with visible and near-infrared excitations. MP4(SDQ) level of theory with the 6-311g(2d,2p) basis set is executed for three different electronic spin states (singlet, triplet, and quintet). In order to simplify the potential energy function, an approximation is adopted by ignoring the spin-orbit coupling and allowing the molecule to switch favorably and instantaneously to the spin state that is more energetically stable (lowest in energy among the three spin states). This assumption has previously been utilized to study the SiO(2) system as reported by Agrawal et al. (J. Chem. Phys. 2006, 124 (13), 134306). The use of such assumption in this study probably makes the upper limits of computed rate coefficients the true rate coefficients. The global PES for ozone is constructed by fitting 5906 ab initio data points using a 60-neuron two-layer feed-forward neural network. The mean-absolute error and root-mean-squared error of this fit are 0.0446 eV (1.03 kcal/mol) and 0.0756 eV (1.74 kcal/mol), respectively, which reveal very good fitting accuracy. The parameter coefficients of the global PES are reported in this paper. In order to identify the spin state with high confidence, we propose the use of a pattern-recognition neural network, which is trained to predict the spin state of a given configuration (with a prediction accuracy being 95.6% on a set of testing data points). To enhance the prediction effectiveness, a buffer series of five points are validated to confirm the spin state during the MD process to gain better confidence. Quasi-classical MD simulations from 1.2 to 2.4 eV of total internal energy (including zero-point energy) result in rate coefficients of singlet-triplet transformation in the range of 0.027 ps(-1) to 1.21 ps(-1). Also, we find very

  15. Molecular recognition and organizational and polyvalent effects in vesicles induce the formation of artificial multicompartment cells as model systems of eukaryotes.

    PubMed

    Paleos, Constantinos M; Pantos, A

    2014-05-20

    Researchers have become increasingly interested in the preparation and characterization of artificial cells based on amphiphilic molecules. In particular, artificial cells with multiple compartments are primitive mimics of the structure of eukaryotic cells. Endosymbiotic theory, widely accepted among biologists, states that eukaryotic cells arose from the assembly of prokaryotic cells inside other cells. Therefore, replicating this process in a synthetic system could allow researchers to model molecular and supramolecular processes that occur in living cells, shed light on mass and energy transport through cell membranes, and provide a unique, isolated space for conducting chemical reactions. In addition, such structures can serve as drug delivery systems that encapsulate both bioactive and nonbiocompatible compounds. In this Account, we present various coating, incubation, and electrofusion strategies for forming multicompartment vesicle systems, and we are focusing on strategies that rely on involving molecular recognition of complementary vesicles. All these methods afforded multicompartment systems with similar structures, and these nanoparticles have potential applications as drug delivery systems or nanoreactors for conducting diverse reactions. The complementarity of interacting vesicles allows these artificial cells to form, and the organization and polyvalency of these interacting vesicles further promote their formation. The incorporation of cholesterol in the bilayer membrane and the introduction of PEG chains at the surface of the interacting vesicles also support the structure of these multicompartment systems. PEG chains appear to destabilize the bilayers, which facilitates the fusion and transport of the small vesicles to the larger ones. Potential applications of these well-structured and reproducibly produced multicompartment systems include drug delivery, where researchers could load a cocktail of drugs within the encapsulated vesicles, a process

  16. Voice recognition.

    PubMed

    Mehta, Amit; McLoud, Theresa C

    2003-07-01

    Voice recognition represents one of the new technologies that are changing the practice of radiology. Thirty percent of radiology practices are either currently or plan to have voice recognition (VR) systems. VR software encompasses 4 core processes: spoken recognition of human speech, synthesis of human readable characters into speech, speaker identification and verification, and comprehension. Many software packages are available offering VR. All these packages should contain an interface with the radiology information system. The benefits include decreased turnaround time and cost savings. Its advantages include the transfer of secretarial duties to the radiologist with a result in decreased productivity. PMID:12867815

  17. Conjugated-protein mimics with molecularly imprinted reconstructible and transformable regions that are assembled using space-filling prosthetic groups.

    PubMed

    Takeuchi, Toshifumi; Mori, Takuya; Kuwahara, Atsushi; Ohta, Takeo; Oshita, Azusa; Sunayama, Hirobumi; Kitayama, Yukiya; Ooya, Tooru

    2014-11-17

    Conjugated-protein mimics were obtained using a new molecular imprinting strategy combined with post-imprinting modifications. An antibiotic was employed as a model template molecule, and a polymerizable template molecule was designed, which was composed of the antibiotic and two different prosthetic groups attached through a disulfide bond and Schiff base formation. After co-polymerization with a cross-linker, the template molecule was removed together with the prosthetic groups, yielding the apo-type scaffold. Through conjugation of the two different prosthetic groups at pre-determined positions within the apo-type scaffold, the apo cavity was transformed into a functionalized holo cavity, which enables the on/off switching of the molecular recognition ability, signal transduction activity for binding events, and photoresponsive activity. PMID:25257234

  18. Instruction, Cognitive Scaffolding, and Motivational Scaffolding in Writing Center Tutoring

    ERIC Educational Resources Information Center

    Mackiewicz, Jo; Thompson, Isabelle

    2014-01-01

    In this study, we quantitatively analyze the discourse of experienced writing center tutors in 10 highly satisfactory conferences. Specifically, we analyze tutors' instruction, cognitive scaffolding, and motivational scaffolding, all tutoring strategies identified in prior research from other disciplines as educationally effective. We find…

  19. Melt electrospinning of biodegradable polyurethane scaffolds

    PubMed Central

    Karchin, Ari; Simonovsky, Felix I.; Ratner, Buddy D.; Sanders, Joan E.

    2014-01-01

    Electrospinning from the melt, in contrast to from solution, is an attractive tissue engineering scaffold manufacturing process as it allows for the formation of small diameter fibers while eliminating potentially cytotoxic solvents. Despite this, there is a dearth of literature on scaffold formation via melt electrospinning. This is likely due to the technical challenges related to the need for a well-controlled high temperature setup and the difficulty in developing an appropriate polymer. In this paper, a biodegradable and thermally stable polyurethane (PU) is described specifically for use in melt electrospinning. Polymer formulations of aliphatic PUs based on (CH2)4-content diisocyanates, polycaprolactone (PCL), 1,4-butanediamine and 1,4-butanediol (BD) were evaluated for utility in the melt electrospinning process. The final polymer formulation, a catalyst-purified PU based on 1,4-butane diisocyanate, PCL and BD in a 4/1/3 molar ratio with a weight-average molecular weight of about 40 kDa, yielded a nontoxic polymer that could be readily electrospun from the melt. Scaffolds electrospun from this polymer contained point bonds between fibers and mechanical properties analogous to many in vivo soft tissues. PMID:21640853

  20. Nucleic acid recognition by tandem helical repeats.

    PubMed

    Rubinson, Emily H; Eichman, Brandt F

    2012-02-01

    Protein domains constructed from tandem α-helical repeats have until recently been primarily associated with protein scaffolds or RNA recognition. Recent crystal structures of human mitochondrial termination factor MTERF1 and Bacillus cereus alkylpurine DNA glycosylase AlkD bound to DNA revealed two new superhelical tandem repeat architectures capable of wrapping around the double helix in unique ways. Unlike DNA sequence recognition motifs that rely mainly on major groove read-out, MTERF and ALK motifs locate target sequences and aberrant nucleotides within DNA by resculpting the double-helix through extensive backbone contacts. Comparisons between MTERF and ALK repeats, together with recent advances in ssRNA recognition by Pumilio/FBF (PUF) domains, provide new insights into the fundamental principles of protein-nucleic acid recognition. PMID:22154606

  1. Speech recognition based on pattern recognition techniques

    NASA Astrophysics Data System (ADS)

    Rabiner, Lawrence R.

    1990-05-01

    Algorithms for speech recognition can be characterized broadly as pattern recognition approaches and acoustic phonetic approaches. To date, the greatest degree of success in speech recognition has been obtained using pattern recognition paradigms. The use of pattern recognition techniques were applied to the problems of isolated word (or discrete utterance) recognition, connected word recognition, and continuous speech recognition. It is shown that understanding (and consequently the resulting recognizer performance) is best to the simplest recognition tasks and is considerably less well developed for large scale recognition systems.

  2. RING domains: master builders of molecular scaffolds?

    PubMed

    Borden, K L

    2000-02-01

    Intense interest in the RING domain has arisen because of its widespread occurrence and involvement in human disease. Several intriguing characteristics evident from the study of this cysteine-rich, zinc-binding domain have made it difficult to establish a single defining biochemical function for RINGs. These proteins are found throughout the cell and mediate diverse cellular processes, e.g. oncogenesis, apoptosis, development and viral infection. Recent developments indicate that RING-mediated protein interactions are critical for transcriptional repression and for ubiquitination. These data are in addition to previously established functions for RINGs in RNA processing, cell-cycle control and peroxisomal biogenesis, to name a few. At first glance, there appears to be little to link such disparate actions. Collectively, these results suggest that RINGs function in formation and architecture of large protein complexes that contribute to diverse cellular processes. Here, new developments, in the context of previous results, are discussed in an attempt to establish a unifying theory for RING function. PMID:10653689

  3. Approaching rational epitope vaccine design for hepatitis C virus with meta-server and multivalent scaffolding

    NASA Astrophysics Data System (ADS)

    He, Linling; Cheng, Yushao; Kong, Leopold; Azadnia, Parisa; Giang, Erick; Kim, Justin; Wood, Malcolm R.; Wilson, Ian A.; Law, Mansun; Zhu, Jiang

    2015-08-01

    Development of a prophylactic vaccine against hepatitis C virus (HCV) has been hampered by the extraordinary viral diversity and the poor host immune response. Scaffolding, by grafting an epitope onto a heterologous protein scaffold, offers a possible solution to epitope vaccine design. In this study, we designed and characterized epitope vaccine antigens for the antigenic sites of HCV envelope glycoproteins E1 (residues 314-324) and E2 (residues 412-423), for which neutralizing antibody-bound structures are available. We first combined six structural alignment algorithms in a “scaffolding meta-server” to search for diverse scaffolds that can structurally accommodate the HCV epitopes. For each antigenic site, ten scaffolds were selected for computational design, and the resulting epitope scaffolds were analyzed using structure-scoring functions and molecular dynamics simulation. We experimentally confirmed that three E1 and five E2 epitope scaffolds bound to their respective neutralizing antibodies, but with different kinetics. We then investigated a “multivalent scaffolding” approach by displaying 24 copies of an epitope scaffold on a self-assembling nanoparticle, which markedly increased the avidity of antibody binding. Our study thus demonstrates the utility of a multi-scale scaffolding strategy in epitope vaccine design and provides promising HCV immunogens for further assessment in vivo.

  4. Approaching rational epitope vaccine design for hepatitis C virus with meta-server and multivalent scaffolding

    PubMed Central

    He, Linling; Cheng, Yushao; Kong, Leopold; Azadnia, Parisa; Giang, Erick; Kim, Justin; Wood, Malcolm R.; Wilson, Ian A.; Law, Mansun; Zhu, Jiang

    2015-01-01

    Development of a prophylactic vaccine against hepatitis C virus (HCV) has been hampered by the extraordinary viral diversity and the poor host immune response. Scaffolding, by grafting an epitope onto a heterologous protein scaffold, offers a possible solution to epitope vaccine design. In this study, we designed and characterized epitope vaccine antigens for the antigenic sites of HCV envelope glycoproteins E1 (residues 314–324) and E2 (residues 412–423), for which neutralizing antibody-bound structures are available. We first combined six structural alignment algorithms in a “scaffolding meta-server” to search for diverse scaffolds that can structurally accommodate the HCV epitopes. For each antigenic site, ten scaffolds were selected for computational design, and the resulting epitope scaffolds were analyzed using structure-scoring functions and molecular dynamics simulation. We experimentally confirmed that three E1 and five E2 epitope scaffolds bound to their respective neutralizing antibodies, but with different kinetics. We then investigated a “multivalent scaffolding” approach by displaying 24 copies of an epitope scaffold on a self-assembling nanoparticle, which markedly increased the avidity of antibody binding. Our study thus demonstrates the utility of a multi-scale scaffolding strategy in epitope vaccine design and provides promising HCV immunogens for further assessment in vivo. PMID:26238798

  5. Macroporous photocrosslinked elastomer scaffolds containing microposity: preparation and in vitro degradation properties.

    PubMed

    Ilagan, Bernadette G; Amsden, Brian G

    2010-04-01

    The engineering of soft tissue would benefit from the development of effective biodegradable scaffolds capable of dynamic, elastic loading. For this purpose, highly porous, elastomeric scaffolds containing microporous struts were prepared using a dual porogen approach and a photocrosslinkable elastomer. The combination of paraffin microbeads distributed through a water-in-[star-poly(lactide-co-epsilon-caprolactone) triacrylate dissolved in ethyl acetate] emulsion followed by photocrosslinking generated a macroporous foam scaffold of average porosities between 90% to 93%, with an average pore diameter of 104 +/- 31 microm with struts containing micropores of 3.1 +/- 2 microm average diameter. The mechanical properties of the scaffolds were readily manipulatable by altering the molecular weight of the star-poly(lactide-co-epsilon-caprolactone) triacrylate prepolymer used. The elastomer scaffolds degraded at the same rate as nonporous polymer samples of the same molecular weight, and exhibited similar changes in mass loss, mechanical properties, and sol fraction during in vitro degradation as found with the nonporous scaffolds. The modulus and stress at break of the scaffolds decreased continuously during degradation while the strain at break remained constant. These scaffolds show potential for use in the engineering of soft tissues. PMID:19544482

  6. Water-stable three-dimensional ultrafine fibrous scaffolds from keratin for cartilage tissue engineering.

    PubMed

    Xu, Helan; Cai, Shaobo; Xu, Lan; Yang, Yiqi

    2014-07-22

    Intrinsically water-stable scaffolds composed of ultrafine keratin fibers oriented randomly and evenly in three dimensions were electrospun for cartilage tissue engineering. Keratin has been recognized as a biomaterial that could substantially support the growth and development of multiple cell lines. Besides, three-dimensional (3D) ultrafine fibrous structures were preferred in tissue engineering due to their structural similarity to native extracellular matrices in soft tissues. Recently, we have developed a nontraditional approach to developing 3D fibrous scaffolds from alcohol-soluble corn protein, zein, and verified their structural advantages in tissue engineering. However, keratin with highly cross-linked molecular structures could not be readily dissolved in common solvents for fiber spinning, which required the remarkable drawability of solution. So far, 3D fibrous scaffolds from pure keratin for biomedical applications have not been reported. In this research, the highly cross-linked keratin from chicken feathers was de-cross-linked and disentangled into linear and aligned molecules with preserved molecular weights, forming highly stretchable spinning dope. The solution was readily electrospun into scaffolds with ultrafine keratin fibers oriented randomly in three dimensions. Due to the highly cross-linked molecular structures, keratin scaffolds showed intrinsic water stability. Adipose-derived mesenchymal stem cells could penetrate much deeper, proliferate, and chondrogenically differentiate remarkably better on the 3D keratin scaffolds than on 2D PLA fibrous scaffolds, 3D soy protein fibrous scaffolds, or 3D commercial nonfibrous scaffolds. In summary, the electrospun 3D ultrafine fibrous scaffolds from keratin could be promising candidates for cartilage tissue engineering. PMID:25010870

  7. Using Scaffolds in Problem-Based Hypermedia

    ERIC Educational Resources Information Center

    Su, Yuyan; Klein, James D.

    2010-01-01

    This study investigated the use of scaffolds in problem-based hypermedia. Three hundred and twelve undergraduate students enrolled in a computer literacy course worked in project teams to use a hypermedia PBL program focused on designing a personal computer. The PBL program included content scaffolds, metacognitive scaffolds, or no scaffolds.…

  8. Electrospun multifunctional tissue engineering scaffolds

    NASA Astrophysics Data System (ADS)

    Wang, Chong; Wang, Min

    2014-03-01

    Tissue engineering holds great promises in providing successful treatments of human body tissue loss that current methods are unable to treat or unable to achieve satisfactory clinical outcomes. In scaffold-based tissue engineering, a highperformance scaffold underpins the success of a tissue engineering strategy and a major direction in the field is to create multifunctional tissue engineering scaffolds for enhanced biological performance and for regenerating complex body tissues. Electrospinning can produce nanofibrous scaffolds that are highly desirable for tissue engineering. The enormous interest in electrospinning and electrospun fibrous structures by the science, engineering and medical communities has led to various developments of the electrospinning technology and wide investigations of electrospun products in many industries, including biomedical engineering, over the past two decades. It is now possible to create novel, multicomponent tissue engineering scaffolds with multiple functions. This article provides a concise review of recent advances in the R & D of electrospun multifunctional tissue engineering scaffolds. It also presents our philosophy and research in the designing and fabrication of electrospun multicomponent scaffolds with multiple functions.

  9. Order versus Disorder: in vivo bone formation within osteoconductive scaffolds.

    PubMed

    Scaglione, Silvia; Giannoni, Paolo; Bianchini, Paolo; Sandri, Monica; Marotta, Roberto; Firpo, Giuseppe; Valbusa, Ugo; Tampieri, Anna; Diaspro, Alberto; Bianco, Paolo; Quarto, Rodolfo

    2012-01-01

    In modern biomaterial design the generation of an environment mimicking some of the extracellular matrix features is envisaged to support molecular cross-talk between cells and scaffolds during tissue formation/remodeling. In bone substitutes chemical biomimesis has been particularly exploited; conversely, the relevance of pre-determined scaffold architecture for regenerated bone outputs is still unclear. Thus we aimed to demonstrate that a different organization of collagen fibers within newly formed bone under unloading conditions can be generated by differently architectured scaffolds. An ordered and confined geometry of hydroxyapatite foams concentrated collagen fibers within the pores, and triggered their self-assembly in a cholesteric-banded pattern, resulting in compact lamellar bone. Conversely, when progenitor cells were loaded onto nanofibrous collagen-based sponges, new collagen fibers were distributed in a nematic phase, resulting mostly in woven isotropic bone. Thus specific biomaterial design relevantly contributes to properly drive collagen fibers assembly to target bone regeneration. PMID:22355786

  10. Design properties of hydrogel tissue-engineering scaffolds

    PubMed Central

    Zhu, Junmin; Marchant, Roger E

    2011-01-01

    This article summarizes the recent progress in the design and synthesis of hydrogels as tissue-engineering scaffolds. Hydrogels are attractive scaffolding materials owing to their highly swollen network structure, ability to encapsulate cells and bioactive molecules, and efficient mass transfer. Various polymers, including natural, synthetic and natural/synthetic hybrid polymers, have been used to make hydrogels via chemical or physical crosslinking. Recently, bioactive synthetic hydrogels have emerged as promising scaffolds because they can provide molecularly tailored biofunctions and adjustable mechanical properties, as well as an extracellular matrix-like microenvironment for cell growth and tissue formation. This article addresses various strategies that have been explored to design synthetic hydrogels with extracellular matrix-mimetic bioactive properties, such as cell adhesion, proteolytic degradation and growth factor-binding. PMID:22026626

  11. Molecular recognition of two 2,4-syn-functionalized (S)-glutamate analogues by the kainate receptor GluK3 ligand binding domain.

    PubMed

    Venskutonytė, Raminta; Larsen, Anja P; Frydenvang, Karla; Gajhede, Michael; Sagot, Emmanuelle; Assaf, Zeinab; Gefflaut, Thierry; Pickering, Darryl S; Bunch, Lennart; Kastrup, Jette S

    2014-10-01

    The kainate receptors are the least studied subfamily of ionotropic glutamate receptors. These receptors are thought to have a neuromodulatory role and have been associated with a variety of disorders in the central nervous system. This makes kainate receptors interesting potential drug targets. Today, structures of the ligand binding domain (LBD) of the kainate receptor GluK3 are only known in complex with the endogenous agonist glutamate, the natural product kainate, and two synthetic agonists. Herein we report structures of GluK3 LBD in complex with two 2,4-syn-functionalized (S)-glutamate analogues to investigate their structural potential as chemical scaffolds. Similar binding affinities at GluK3 were determined for the 2-(methylcarbamoyl)ethyl analogue (Ki =4.0 μM) and the 2-(methoxycarbonyl)ethyl analogue (Ki =1.7 μM), in agreement with the similar positioning of the compounds within the binding pocket. As the binding affinity is similar to that of glutamate, this type of Cγ substituent could be used as a scaffold for introduction of even larger substituents reaching into unexplored binding site regions to achieve subtype selectivity. PMID:25044437

  12. Peripheral nerve morphogenesis induced by scaffold micropatterning.

    PubMed

    Cerri, Federica; Salvatore, Luca; Memon, Danish; Martinelli Boneschi, Filippo; Madaghiele, Marta; Brambilla, Paola; Del Carro, Ubaldo; Taveggia, Carla; Riva, Nilo; Trimarco, Amelia; Lopez, Ignazio D; Comi, Giancarlo; Pluchino, Stefano; Martino, Gianvito; Sannino, Alessandro; Quattrini, Angelo

    2014-04-01

    Several bioengineering approaches have been proposed for peripheral nervous system repair, with limited results and still open questions about the underlying molecular mechanisms. We assessed the biological processes that occur after the implantation of collagen scaffold with a peculiar porous micro-structure of the wall in a rat sciatic nerve transection model compared to commercial collagen conduits and nerve crush injury using functional, histological and genome wide analyses. We demonstrated that within 60 days, our conduit had been completely substituted by a normal nerve. Gene expression analysis documented a precise sequential regulation of known genes involved in angiogenesis, Schwann cells/axons interactions and myelination, together with a selective modulation of key biological pathways for nerve morphogenesis induced by porous matrices. These data suggest that the scaffold's micro-structure profoundly influences cell behaviors and creates an instructive micro-environment to enhance nerve morphogenesis that can be exploited to improve recovery and understand the molecular differences between repair and regeneration. PMID:24559639

  13. Quenching of fluorescein-conjugated lipids by antibodies. Quantitative recognition and binding of lipid-bound haptens in biomembrane models, formation of two-dimensional protein domains and molecular dynamics simulations.

    PubMed Central

    Ahlers, M; Grainger, D W; Herron, J N; Lim, K; Ringsdorf, H; Salesse, C

    1992-01-01

    Three model biomembrane systems, monolayers, micelles, and vesicles, have been used to study the influence of chemical and physical variables of hapten presentation at membrane interfaces on antibody binding. Hapten recognition and binding were monitored for the anti-fluorescein monoclonal antibody 4-4-20 generated against the hapten, fluorescein, in these membrane models as a function of fluorescein-conjugated lipid architecture. Specific recognition and binding in this system are conveniently monitored by quenching of fluorescein emission upon penetration of fluorescein into the antibody's active site. Lipid structure was shown to play a large role in affecting antibody quenching. Interestingly, the observed degrees of quenching were nearly independent of the lipid membrane model studied, but directly correlated with the chemical structure of the lipids. In all cases, the antibody recognized and quenched most efficiently a lipid based on dioctadecylamine where fluorescein is attached to the headgroup via a long, flexible hydrophilic spacer. Dipalmitoyl phosphatidylethanolamine containing a fluorescein headgroup demonstrated only partial binding/quenching. Egg phosphatidylethanolamine with a fluorescein headgroup showed no susceptibility to antibody recognition, binding, or quenching. Formation of two-dimensional protein domains upon antibody binding to the fluorescein-lipids in monolayers is also presented. Chemical and physical requirements for these antibody-hapten complexes at membrane surfaces have been discussed in terms of molecular dynamics simulations based on recent crystallographic models for this antibody-hapten complex (Herron et al., 1989. Proteins Struct. Funct. Genet. 5:271-280). Images FIGURE 7 FIGURE 8 PMID:1420916

  14. Cesium cation templated selective synthesis of a "cone-shaped" sugar macrotricyclic cryptand: A dual anion-cation molecular recognition of potassium tartrate.

    PubMed

    Porwanski, S; Moretti, F; Dumarcay-Charbonnier, F; Marsura, A

    2016-05-01

    Cesium templated Staudinger-aza-Wittig tandem reaction (S.A.W.) has been used in the synthesis of a bis-diazacrown-bis-cellobiosyl-tetra-ureido cryptand. A novel macrotricyclic compound having a "cone-shaped" configuration was selectively obtained. Additionally, first results on potential recognition properties of the cryptand are also given. PMID:26826794

  15. Scaffold Diversity Synthesis and Its Application in Probe and Drug Discovery.

    PubMed

    Garcia-Castro, Miguel; Zimmermann, Stefan; Sankar, Muthukumar G; Kumar, Kamal

    2016-06-27

    Scaffold diversity is a crucial feature of compound collections that has a huge impact on their success in biological screenings. The synthesis of highly complex and diverse scaffolds, which could be based on natural products, for example, is an arduous task that requires expertise in various aspects of organic synthesis and structural analysis. This challenge has been addressed by a number of synthesis designs, which employ natural products as a source of scaffold diversity, transform suitably designed common intermediates into various molecular frameworks, or entail highly concise synthetic routes to a number of distinct and complex scaffolds. In this Minireview, we highlight recent synthetic developments towards the construction of diverse and complex scaffolds and the application of the resulting compound collections in drug and probe discovery. PMID:27187638

  16. Peripheral nerve morphogenesis induced by scaffold micropatterning

    PubMed Central

    Memon, Danish; Boneschi, Filippo Martinelli; Madaghiele, Marta; Brambilla, Paola; Del Carro, Ubaldo; Taveggia, Carla; Riva, Nilo; Trimarco, Amelia; Lopez, Ignazio D.; Comi, Giancarlo; Pluchino, Stefano; Martino, Gianvito; Sannino, Alessandro; Quattrini, Angelo

    2014-01-01

    Several bioengineering approaches have been proposed for peripheral nervous system repair, with limited results and still open questions about the underlying molecular mechanisms. We assessed the biological processes that occur after the implantation of collagen scaffold with a peculiar porous microstructure of the wall in a rat sciatic nerve transection model compared to commercial collagen conduits and nerve crush injury using functional, histological and genome wide analyses. We demonstrated that within 60 days, our conduit had been completely substituted by a normal nerve. Gene expression analysis documented a precise sequential regulation of known genes involved in angiogenesis, Schwann cells/axons interactions and myelination, together with a selective modulation of key biological pathways for nerve morphogenesis induced by porous matrices. These data suggest that the scaffold’s microstructure profoundly influences cell behaviors and creates an instructive micro-environment to enhance nerve morphogenesis that can be exploited to improve recovery and understand the molecular differences between repair and regeneration. PMID:24559639

  17. Nanotechnology Biomimetic Cartilage Regenerative Scaffolds

    PubMed Central

    Sardinha, Jose Paulo; Myers, Simon

    2014-01-01

    Cartilage has a limited regenerative capacity. Faced with the clinical challenge of reconstruction of cartilage defects, the field of cartilage engineering has evolved. This article reviews current concepts and strategies in cartilage engineering with an emphasis on the application of nanotechnology in the production of biomimetic cartilage regenerative scaffolds. The structural architecture and composition of the cartilage extracellular matrix and the evolution of tissue engineering concepts and scaffold technology over the last two decades are outlined. Current advances in biomimetic techniques to produce nanoscaled fibrous scaffolds, together with innovative methods to improve scaffold biofunctionality with bioactive cues are highlighted. To date, the majority of research into cartilage regeneration has been focused on articular cartilage due to the high prevalence of large joint osteoarthritis in an increasingly aging population. Nevertheless, the principles and advances are applicable to cartilage engineering for plastic and reconstructive surgery. PMID:24883273

  18. Multilayered Magnetic Gelatin Membrane Scaffolds.

    PubMed

    Samal, Sangram K; Goranov, Vitaly; Dash, Mamoni; Russo, Alessandro; Shelyakova, Tatiana; Graziosi, Patrizio; Lungaro, Lisa; Riminucci, Alberto; Uhlarz, Marc; Bañobre-López, Manuel; Rivas, Jose; Herrmannsdörfer, Thomas; Rajadas, Jayakumar; De Smedt, Stefaan; Braeckmans, Kevin; Kaplan, David L; Dediu, V Alek

    2015-10-21

    A versatile approach for the design and fabrication of multilayer magnetic scaffolds with tunable magnetic gradients is described. Multilayer magnetic gelatin membrane scaffolds with intrinsic magnetic gradients were designed to encapsulate magnetized bioagents under an externally applied magnetic field for use in magnetic-field-assisted tissue engineering. The temperature of the individual membranes increased up to 43.7 °C under an applied oscillating magnetic field for 70 s by magnetic hyperthermia, enabling the possibility of inducing a thermal gradient inside the final 3D multilayer magnetic scaffolds. On the basis of finite element method simulations, magnetic gelatin membranes with different concentrations of magnetic nanoparticles were assembled into 3D multilayered scaffolds. A magnetic-gradient-controlled distribution of magnetically labeled stem cells was demonstrated in vitro. This magnetic biomaterial-magnetic cell strategy can be expanded to a number of different magnetic biomaterials for various tissue engineering applications. PMID:26451743

  19. Halogen bonding anion recognition.

    PubMed

    Brown, Asha; Beer, Paul D

    2016-07-01

    A halogen bond is an attractive non-covalent interaction between an electrophilic region in a covalently bonded halogen atom and a Lewis base. While these interactions have long been exploited as a tool in crystal engineering their powerful ability to direct supramolecular self-assembly and molecular recognition processes in solution has, until recently, been overlooked. During the last decade however an ever-increasing number of studies on solution-phase halogen-bond-mediated anion recognition processes has emerged. This Feature Article summarises advancements which have been made thus far in this rapidly developing research area. We survey the use of iodoperfluoroarene, haloimidazolium and halotriazole/triazolium halogen-bond-donor motifs in anion receptor design, before providing an account of our research into the application of mechanically interlocked rotaxane and catenane frameworks as halogen bonding anion host systems. PMID:27273600

  20. [Articular cartilage regeneration using scaffold].

    PubMed

    Ishimoto, Yoshiyuki; Hattori, Koji; Ohgushi, Hajime

    2008-12-01

    The self-healing capacity of articular cartilage for repair is limited. For articular cartilage injury, several surgical techniques are used in clinical practice, namely drilling, abrasion arthroplasty, microfracture, or autologous osteochondral grafting, while various methods of autologous chondrocyte transplantation to cartilage defect sites have been reported since 1990s. In a case of chondrocyte transplantation to cartilage defect site, the use of proper scaffold is important. Currently, collagen gel or PLGA is used widely as a scaffold. PMID:19043192

  1. Heterocyclic Scaffolds in the Design of Peptidomimetic Integrin Ligands: Synthetic Strategies, Structural Aspects, and Biological Activity.

    PubMed

    De Marco, Rossella; Mazzotti, Giacomo; Greco, Arianna; Gentilucci, Luca

    2016-01-01

    The integrin receptors represent valuable targets for therapeutic interventions; being overexpressed in many pathological states, their inhibition can be effective to treat a number of severe diseases. Since integrin functions are mediated by interactions with ECM protein ligands, the inhibition can be achieved by interfering with such interactions using small mimetics of the integrin-ligand recognition motifs (e.g. RGD, LDV, etc.). In this review, we focus on the antagonists with peptideheterocycle hybrid structures. The introduction of well-designed scaffolds has met considerable success in the rational design of highly stable, bioavailable, and conformationally defined antagonists. Two main approaches are discussed herein. The first approach is the use of scaffolds external to the main recognition motifs, aimed at improving conformational definition. In the second approach, heterocyclic cores are introduced within the recognition motifs, giving access to libraries of 3D diverse candidate antagonists. PMID:26265351

  2. Programmable DNA scaffolds for spatially-ordered protein assembly

    NASA Astrophysics Data System (ADS)

    Chandrasekaran, Arun Richard

    2016-02-01

    Ever since the notion of using DNA as a material was realized, it has been employed in the construction of complex structures that facilitate the assembly of nanoparticles or macromolecules with nanometer-scale precision. Specifically, tiles fashioned from DNA strands and DNA origami sheets have been shown to be suitable as scaffolds for immobilizing proteins with excellent control over their spatial positioning. Supramolecular assembly of proteins into periodic arrays in one or more dimensions is one of the most challenging aspects in the design of scaffolds for biomolecular investigations and macromolecular crystallization. This review provides a brief overview of how various biomolecular interactions with high degree of specificity such as streptavidin-biotin, antigen-antibody, and aptamer-protein interactions have been used to fabricate linear and multidimensional assemblies of structurally intact and functional proteins. The use of DNA-binding proteins as adaptors, polyamide recognition on DNA scaffolds and oligonucleotide linkers for protein assembly are also discussed.Ever since the notion of using DNA as a material was realized, it has been employed in the construction of complex structures that facilitate the assembly of nanoparticles or macromolecules with nanometer-scale precision. Specifically, tiles fashioned from DNA strands and DNA origami sheets have been shown to be suitable as scaffolds for immobilizing proteins with excellent control over their spatial positioning. Supramolecular assembly of proteins into periodic arrays in one or more dimensions is one of the most challenging aspects in the design of scaffolds for biomolecular investigations and macromolecular crystallization. This review provides a brief overview of how various biomolecular interactions with high degree of specificity such as streptavidin-biotin, antigen-antibody, and aptamer-protein interactions have been used to fabricate linear and multidimensional assemblies of structurally

  3. A mixture approach to investigate interstitial growth in engineering scaffolds.

    PubMed

    Vernerey, Franck J

    2016-04-01

    Controlling biological growth within a cell-laden polymeric scaffold is a critical challenge in the tissue engineering community. Indeed, construct growth must often be balanced with scaffold degradation and is often coupled to varying degrees of deformation that originate from swelling, external forces and the effects of confinement. These factors have been shown to affect growth in many ways, but to date, our understanding is mostly qualitative. While cell sensing, molecular transport and scaffold/tissue interactions are believed to be important players, it will be critical to quantify, predict and control these effects in order to eventually optimize tissue growth in the laboratory. The aim of this paper was thus to provide a theoretical framework to better understand how the scaffold-mediated mechanisms of transport, deposition (and possibly degradation) and elasticity affect the overall growth of a tissue subjected to finite deformations. We propose a formulation in which the macroscopic evolutions in tissue size, density as well as the appearance of residual stresses can be directly related to changes in internal composition by considering three fundamental principles: mechanical equilibrium, chemical equilibrium and molecular incompressibility. The resulting model allows us to pay particular attention to features that are critical to the interaction between growth and deformation: osmotic pressure and swelling, the strain mismatch between old and newly deposited material as well as the mechano-sensitive cell-mediated production. We show that all of these phenomena may indeed strongly affect the overall growth of a construct under finite deformations. PMID:26047777

  4. Polyurethane/poly(lactic-co-glycolic) acid composite scaffolds fabricated by thermally induced phase separation.

    PubMed

    Rowlands, A S; Lim, S A; Martin, D; Cooper-White, J J

    2007-04-01

    In this study, we present a novel composite scaffold fabricated using a thermally induced phase separation (TIPS) process from poly(lactic-co-glycolic) (PLGA) and biomedical polyurethane (PU). This processing method has been tuned to allow intimate (molecular) mixing of these two very different polymers, giving rise to a unique morphology that can be manipulated by controlling the phase separation behaviour of an initially homogenous polymer solution. Pure PLGA scaffolds possessed a smooth, directional fibrous sheet-like structure with pore sizes of 0.1-200mum, a porous Young's modulus of 93.5kPa and were relatively brittle to touch. Pure PU scaffolds had an isotropic emulsion-like structure, a porous Young's modulus of 15.7kPa and were much more elastic than the PLGA scaffolds. The composite PLGA/PU scaffold exhibits advantageous morphological, mechanical and cell adhesion and growth supporting properties, when compared with scaffolds fabricated from PLGA or PU alone. This novel method provides a mechanism for the formation of tailored bioactive scaffolds from nominally incompatible polymers, representing a significant step forward in scaffold processing for tissue-engineering applications. PMID:17258315

  5. Experimental and Modeling Study of Collagen Scaffolds with the Effects of Crosslinking and Fiber Alignment

    PubMed Central

    Xu, Bin; Chow, Ming-Jay; Zhang, Yanhang

    2011-01-01

    Collagen type I scaffolds are commonly used due to its abundance, biocompatibility, and ubiquity. Most applications require the scaffolds to operate under mechanical stresses. Therefore understanding and being able to control the structural-functional integrity of collagen scaffolds becomes crucial. Using a combined experimental and modeling approach, we studied the structure and function of Type I collagen gel with the effects of spatial fiber alignment and crosslinking. Aligned collagen scaffolds were created through the flow of magnetic particles enmeshed in collagen fibrils to mimic the anisotropy seen in native tissue. Inter- and intra- molecular crosslinking was modified chemically with Genipin to further improve the stiffness of collagen scaffolds. The anisotropic mechanical properties of collagen scaffolds were characterized using a planar biaxial tensile tester and parallel plate rheometer. The tangent stiffness from biaxial tensile test is two to three orders of magnitude higher than the storage moduli from rheological measurements. The biphasic nature of collagen gel was discussed and used to explain the mechanical behavior of collagen scaffolds under different types of mechanical tests. An anisotropic hyperelastic constitutive model was used to capture the characteristics of the stress-strain behavior exhibited by collagen scaffolds. PMID:21876695

  6. Electrospun silk fibroin/poly(lactide-co-ε-caprolactone) nanofibrous scaffolds for bone regeneration

    PubMed Central

    Wang, Zi; Lin, Ming; Xie, Qing; Sun, Hao; Huang, Yazhuo; Zhang, DanDan; Yu, Zhang; Bi, Xiaoping; Chen, Junzhao; Wang, Jing; Shi, Wodong; Gu, Ping; Fan, Xianqun

    2016-01-01

    Background Tissue engineering has become a promising therapeutic approach for bone regeneration. Nanofibrous scaffolds have attracted great interest mainly due to their structural similarity to natural extracellular matrix (ECM). Poly(lactide-co-ε-caprolactone) (PLCL) has been successfully used in bone regeneration, but PLCL polymers are inert and lack natural cell recognition sites, and the surface of PLCL scaffold is hydrophobic. Silk fibroin (SF) is a kind of natural polymer with inherent bioactivity, and supports mesenchymal stem cell attachment, osteogenesis, and ECM deposition. Therefore, we fabricated hybrid nanofibrous scaffolds by adding different weight ratios of SF to PLCL in order to find a scaffold with improved properties for bone regeneration. Methods Hybrid nanofibrous scaffolds were fabricated by blending different weight ratios of SF with PLCL. Human adipose-derived stem cells (hADSCs) were seeded on SF/PLCL nanofibrous scaffolds of various ratios for a systematic evaluation of cell adhesion, proliferation, cytotoxicity, and osteogenic differentiation; the efficacy of the composite of hADSCs and scaffolds in repairing critical-sized calvarial defects in rats was investigated. Results The SF/PLCL (50/50) scaffold exhibited favorable tensile strength, surface roughness, and hydrophilicity, which facilitated cell adhesion and proliferation. Moreover, the SF/PLCL (50/50) scaffold promoted the osteogenic differentiation of hADSCs by elevating the expression levels of osteogenic marker genes such as BSP, Ocn, Col1A1, and OPN and enhanced ECM mineralization. In vivo assays showed that SF/PLCL (50/50) scaffold improved the repair of the critical-sized calvarial defect in rats, resulting in increased bone volume, higher trabecular number, enhanced bone mineral density, and increased new bone areas, compared with the pure PLCL scaffold. Conclusion The SF/PLCL (50/50) nanofibrous scaffold facilitated hADSC proliferation and osteogenic differentiation in

  7. Polymer-based separations: Synthesis and application of polymers for ionic and molecular recognition. Triennial performance report, August 1, 1989--July 31, 1992

    SciTech Connect

    Alexandratos, S.D.

    1992-01-01

    Polymer-based separations have utilized resins such as sulfonic, acrylic, and iminodiacetic acid resins and the XAD series. Selective polymeric reagents for reaction with a targeted metal ion were synthesized as polymers with two different types of functional groups, each operating on the ions through a different mechanism. There are 3 classes of DMBPs (dual mechanism bifunctional polymers). Research during this period dealing with metal ion recognition focused on two of these classes (reduction of metal ions to metal; selective complexation).

  8. Teaching an Old Scaffold New Tricks: Monobodies Constructed Using Alternative Surfaces of the FN3 Scaffold

    SciTech Connect

    Koide, Akiko; Wojcik, John; Gilbreth, Ryan N.; Hoey, Robert J.; Koide, Shohei

    2012-06-28

    The fibronectin type III domain (FN3) has become one of the most widely used non-antibody scaffolds for generating new binding proteins. Because of its structural homology to the immunoglobulin domain, combinatorial libraries of FN3 designed to date have primarily focused on introducing amino acid diversity into three loops that are equivalent to antibody complementarity-determining regions. Here, we report an FN3 library that utilizes alternative positions for presenting amino acid diversity. We diversified positions on a {beta}-sheet and surface loops that together form a concave surface. The new library produced binding proteins (termed 'monobodies') to multiple target proteins, generally with similar efficacy as the original, loop-focused library. The crystal structure of a monobody generated from the new library in complex with its target, the Abl SH2 domain, revealed that a concave surface of the monobody, as intended in our design, bound to a convex surface of the target with the interface area being among the largest of published structures of monobody-target complexes. This mode of interaction differs from a common binding mode for single-domain antibodies and antibody mimics in which recognition loops recognize clefts in targets. Together, this work illustrates the utilization of different surfaces of a single immunoglobulin-like scaffold to generate binding proteins with distinct characteristics.

  9. Laser microstructured biodegradable scaffolds.

    PubMed

    Koroleva, Anastasia; Kufelt, Olga; Schlie-Wolter, Sabrina; Hinze, Ulf; Chichkov, Boris

    2013-10-01

    The two-photon polymerization technique (2PP) uses non-linear absorption of femtosecond laser pulses to selectively polymerize photosensitive materials. 2PP has the ability to fabricate structures with a resolution from tens of micrometers down to hundreds of nanometers. Three-dimensional microstructuring by the 2PP technique provides many interesting possibilities for biomedical applications. This microstructuring technique is suitable with many biocompatible polymeric materials, such as polyethylene glycol, polylactic acid, polycaprolactone, gelatin, zirconium-based hybrids, and others. The process of fabrication does not require clean room conditions and does not use hazard chemicals or high temperatures. The most beneficial property of 2PP is that it is capable of producing especially complex three-dimensional (3-D) structures, including devices with overhangs, without using any supportive structure. The flexibility in controlling geometries and feature sizes and the possibility to fabricate structures without the addition of new material layers makes this technique particularly appealing for fabrication of 3-D scaffolds for tissue engineering. PMID:23729598

  10. Neuronal Networks on Nanocellulose Scaffolds.

    PubMed

    Jonsson, Malin; Brackmann, Christian; Puchades, Maja; Brattås, Karoline; Ewing, Andrew; Gatenholm, Paul; Enejder, Annika

    2015-11-01

    Proliferation, integration, and neurite extension of PC12 cells, a widely used culture model for cholinergic neurons, were studied in nanocellulose scaffolds biosynthesized by Gluconacetobacter xylinus to allow a three-dimensional (3D) extension of neurites better mimicking neuronal networks in tissue. The interaction with control scaffolds was compared with cationized nanocellulose (trimethyl ammonium betahydroxy propyl [TMAHP] cellulose) to investigate the impact of surface charges on the cell interaction mechanisms. Furthermore, coatings with extracellular matrix proteins (collagen, fibronectin, and laminin) were investigated to determine the importance of integrin-mediated cell attachment. Cell proliferation was evaluated by a cellular proliferation assay, while cell integration and neurite propagation were studied by simultaneous label-free Coherent anti-Stokes Raman Scattering and second harmonic generation microscopy, providing 3D images of PC12 cells and arrangement of nanocellulose fibrils, respectively. Cell attachment and proliferation were enhanced by TMAHP modification, but not by protein coating. Protein coating instead promoted active interaction between the cells and the scaffold, hence lateral cell migration and integration. Irrespective of surface modification, deepest cell integration measured was one to two cell layers, whereas neurites have a capacity to integrate deeper than the cell bodies in the scaffold due to their fine dimensions and amoeba-like migration pattern. Neurites with lengths of >50 μm were observed, successfully connecting individual cells and cell clusters. In conclusion, TMAHP-modified nanocellulose scaffolds promote initial cellular scaffold adhesion, which combined with additional cell-scaffold treatments enables further formation of 3D neuronal networks. PMID:26398224

  11. Bilayer membrane interactions with nanofabricated scaffolds

    DOE PAGESBeta

    Collier, C. Patrick

    2015-07-29

    Membrane function is facilitated by lateral organization within the lipid bilayer, including phase-separation of lipids into more ordered domains (lipid rafts) and anchoring of the membrane to a cytoskeleton. These features have proven difficult to reproduce in model membrane systems such as black lipid membranes, unilamellar vesicles and supported bilayers. However, advances in micro/nanofabrication have resulted in more realistic synthetic models of membrane-cytoskeleton interactions that can help uncover the design rules responsible for biological membrane formation and organization. This review will focus on describing micro-/nanostructured scaffolds that can emulate the connections of a cellular membrane to an underlying “cytoskeleton”. Thismore » includes molecular-based scaffolds anchored to a solid substrate through surface chemistry, solid-state supports modified by material deposition, lithography and etching, the creation of micro/nanoporous arrays, integration with microfluidics, and droplet-based bilayers at interfaces. Lastly, model systems such as these are increasing our understanding of structure and organization in cell membranes, and how they result in the emergence of functionality at the nanoscale.« less

  12. Bilayer membrane interactions with nanofabricated scaffolds

    SciTech Connect

    Collier, C. Patrick

    2015-07-29

    Membrane function is facilitated by lateral organization within the lipid bilayer, including phase-separation of lipids into more ordered domains (lipid rafts) and anchoring of the membrane to a cytoskeleton. These features have proven difficult to reproduce in model membrane systems such as black lipid membranes, unilamellar vesicles and supported bilayers. However, advances in micro/nanofabrication have resulted in more realistic synthetic models of membrane-cytoskeleton interactions that can help uncover the design rules responsible for biological membrane formation and organization. This review will focus on describing micro-/nanostructured scaffolds that can emulate the connections of a cellular membrane to an underlying “cytoskeleton”. This includes molecular-based scaffolds anchored to a solid substrate through surface chemistry, solid-state supports modified by material deposition, lithography and etching, the creation of micro/nanoporous arrays, integration with microfluidics, and droplet-based bilayers at interfaces. Lastly, model systems such as these are increasing our understanding of structure and organization in cell membranes, and how they result in the emergence of functionality at the nanoscale.

  13. Osteogenic Scaffolds for Bone Reconstruction

    PubMed Central

    Li, Ling-jiang; Liu, Ning; Liu, Qing; Jia, Lian-shun; Yuan, Wen

    2012-01-01

    Abstract A highly osteogenic hybrid bioabsorbable scaffold was developed for bone reconstruction/augmentation. Through the use of a solid free-form fabrication technology, a bioabsorbable polycaprolactone (PCL) cage scaffold with a desired size and shape was produced and then filled with osteogenic bone graft particles, that is, morselized autologous bone chips. A rabbit total lamina defect model was chosen to demonstrate its efficacy in regenerating bone with a complicated anatomic shape. Both iliac bone and morselized iliac bone grafts were used in this study for comparison purposes. Serum osteocalcin and collagen type I cross-linked C-terminal telopeptide (CTx) determination showed that active bone remodeling occurred after bone grafts were implanted. X-ray images showed that the bony defects were completely filled with bone mass in all the groups with bone grafts. However, biomechanical tests showed that only the iliac bone and hybrid scaffold groups could restore the mechanical properties to the normal level after 10 weeks of implantation. A histology study showed that both iliac and hybrid scaffold groups had extensive new bone formation, and no adhesion and fibrosis were found. These results indicated that this osteogenic hybrid scaffold can be a good alternative to autologous iliac bone, because it does not need a second iliac bone-harvesting surgery, and thus the morbidity and the possible infections that are often associated with the bone harvesting surgery can be avoided. PMID:23515416

  14. Engineering microporosity in bacterial cellulose scaffolds.

    PubMed

    Bäckdahl, Henrik; Esguerra, Maricris; Delbro, Dick; Risberg, Bo; Gatenholm, Paul

    2008-08-01

    The scaffold is an essential component in tissue engineering. A novel method to prepare three-dimensional (3D) nanofibril network scaffolds with controlled microporosity has been developed. By placing paraffin wax and starch particles of various sizes in a growing culture of Acetobacter xylinum, bacterial cellulose scaffolds of different morphologies and interconnectivity were prepared. Paraffin particles were incorporated throughout the scaffold, while starch particles were found only in the outermost area of the resulting scaffold. The porogens were successfully removed after culture with bacteria and no residues were detected with electron spectroscopy for chemical analysis (ESCA) or Fourier transform infra-red spectroscopy (FT-IR). Resulting scaffolds were seeded with smooth muscle cells (SMCs) and investigated using histology and organ bath techniques. SMC were selected as the cell type since the main purpose of the resulting scaffolds is for tissue engineered blood vessels. SMCs attached to and proliferated on and partly into the scaffolds. PMID:18615821

  15. MAP kinase cascades: scaffolding signal specificity.

    PubMed

    van Drogen, Frank; Peter, Matthias

    2002-01-22

    Scaffold proteins organize many MAP kinase pathways by interacting with several components of these cascades. Recent studies suggest that scaffold proteins provide local activation platforms that contribute to signal specificity by insulating different MAP kinase pathways. PMID:11818078

  16. Recognition intent and visual word recognition.

    PubMed

    Wang, Man-Ying; Ching, Chi-Le

    2009-03-01

    This study adopted a change detection task to investigate whether and how recognition intent affects the construction of orthographic representation in visual word recognition. Chinese readers (Experiment 1-1) and nonreaders (Experiment 1-2) detected color changes in radical components of Chinese characters. Explicit recognition demand was imposed in Experiment 2 by an additional recognition task. When the recognition was implicit, a bias favoring the radical location informative of character identity was found in Chinese readers (Experiment 1-1), but not nonreaders (Experiment 1-2). With explicit recognition demands, the effect of radical location interacted with radical function and word frequency (Experiment 2). An estimate of identification performance under implicit recognition was derived in Experiment 3. These findings reflect the joint influence of recognition intent and orthographic regularity in shaping readers' orthographic representation. The implication for the role of visual attention in word recognition was also discussed. PMID:19036609

  17. The structure of YqeH: An AtNOS1/AtNOA1 ortholog that couples GTP hydrolysis to molecular recognition

    SciTech Connect

    Sudhamsu, J.; Lee, G.I.; Klessig, D.F.; Crane, B.R.

    2009-03-27

    AtNOS1/AtNOA1 was identified as a nitric oxide-generating enzyme in plants, but that function has recently been questioned. To resolve issues surrounding AtNOA1 activity, we report the biochemical properties and a 2.36 {angstrom} resolution crystal structure of a bacterial AtNOA1 ortholog (YqeH). Geobacillus YqeH fused to a putative AtNOA1 leader peptide complements growth and morphological defects of Atnoa1 mutant plants. YqeH does not synthesize nitric oxide from L-arginine but rather hydrolyzes GTP. The YqeH structure reveals a circularly permuted GTPase domain and an unusual C-terminal {beta}-domain. A small N-terminal domain, disordered in the structure, binds zinc. Structural homology among the C-terminal domain, the RNA-binding regulator TRAP, and the hypoxia factor pVHL define a recognition module for peptides and nucleic acids. TRAP residues important for RNA binding are conserved by the YqeH C-terminal domain, whose positioning is coupled to GTP hydrolysis. YqeH and AtNOA1 probably act as G-proteins that regulate nucleic acid recognition and n