M. Kamath

Molecular recognition between a biotinylated polythiophene copolymer and phycoerythrin utilizing the biotin-streptavidin interaction

Abstract The interaction of photoactive proteins with an electroactive matrix is valuable in leading to potential technological applications in opto-electronic signal transduction for optical displays, color mimicking, biosensor, and protein research applications. In this study, the Langmuir-Blodgett technique is used to couple these components through the well-known biotin-streptavidin complexation. Biotinylated copolymers of 3-substituted thiophenes have been synthesized both to enhance mechanical film integrity and to provide the electroactive matrix for attachment of the antennae protein, phycoerythrin. It has been determined that biotinylation of the thiopene copolymers improves their film forming properties and results in stable monolayers. Pressure-area isotherms indicate that protein interaction with the polymer monolayer is occuring with the streptavidin and phycoerythrin proteins. Fluorescene spectroscopy of transferred films confirms the presence of phycoerythrin in the final molecular assemblies. Each stage of protein binding to the biotinylated copolymer monolayer could be monitored through fluorescence microscopy at the air-water interface. These results suggest that this copolymer system is a promising material for integrating virtually any biotinylated macromolecular system to an electroactive matrix. In addition, this copolymer system may be “fine tuned” to maximize protein integration by varying the distance between biotin functionalities and/or the length of the biotin spacer arm.

Intelligent Materials Properties of DNA and Strategies for Its Incorporation into Electroactive Polymeric Thin Film Systems

We propose to create a novel class of intelligent materials by integrating two separate classes of intelligent materials—one biological and the other a thin film conducting polymer. The first class, DNA possesses superior intelligent material properties designed over evolutionary time to function specifically and efficiently in integrated macromolecular arrays in cells called chromo somes. The second material is the polymeric thin film or two-dimensional Langmuir Blodgett (LB) monolayer film. In our approach, films will be comprised of electroactive alkylated conducting polymeric materials, such as polyalkylpyrrole and polyalkylthiophene, that are derivatized with biotin. Steptavidin conjugated DNA will be attached directly or biotinylated DNA will be stably at tached to this film via a bridging streptavidin protein. To date, the bulk of our work has centered on characterizing the DNA binding to thick films of conducting polymers. A near term aim is to incor porate this signal transduction system into fiber optic biosensors for specifically detecting nucleic acid analyte. Our ultimate aim is to create novel ordered structures possessing unique integrated in telligent functions which respond to their environment and provide signal transduction approaches via their electronic and optical functions.

Cerenkov type phase-matched second harmonic generation in polymeric channel waveguides

Abstract Fabrication of polymer channel waveguides by etching on glass is reported. Results on second harmonic generation (SHG) in the Cerenkov mode from stable poled polymeric channel waveguides are presented. The waveguide conversion efficiency for doubling of Nd:YAG laser beam was estimated to be 0.1%/W with a total efficiency of 4.9×10 −7 %/W.

Biotinylated Poly(3-Hexylthiophene-co-3-Methanolthiophene): A Langmuir Monolayer-Forming Copolymer

Abstract Copolymers of 3-substituted thiophenes have been synthesized by organosynthetic routes. The chemical synthesis of the copolymer was carried out by dehydrogenation of 3-hexylthiophene and 3-methanolthiophene. Attachment of biotin to the resulting copolymer, poly(3-hexylthiophene-co-3-methanolthiophene) [PMHT], is accomplished by room temperature esterification using N,N-dicyclohexylcarbodiimide (DCC) and 4-pyrrolidinopyridine as catalyst. The resulting copolymers have well-defined chemical and electronic structures and molecular weights. The biotinylated copolymer forms a stable monolayer at the air-water interface due to the polar groups along the polymer backbone.

Intelligent Systems Based on Ordered Arrays of Biological Molecules Using the LB Technique

The Langmuir-Blodgett (LB) technique has been used to incorporate photodynamic proteins into molecular assemblies. One technique of incorporation involves a generalized cassette attachment methodology which employs a biotin-streptavidin complexation with an electroactive polymer matrix. We have focused the efforts described here on phycobiliproteins and bacteriorho dopsin, although antibodies, enzymes, gene probes and other moieties could also be coupled into the system to build in selectivity. Photoconductivity and photobleaching of these protein systems were investigated. These results suggest that coupling these proteins, either in mixed monolayers or in multilayers, with appropriate conductive polymers or other materials will provide the opto electronic signal transduction needed for biosensor, optical display and other applications. Initial studies involving the integration of conductive polymers into the molecular assemblies to enhance optical signal transduction are also discussed.

Second-order nonlinear optical properties of a polymer exhibiting optical transparency down to 340 nm

Abstract The synthesis and characterization of an epoxy-based nonlinear optical (NLO) polymer exhibiting optical transparency down to 340 nm is reported. The synthesized polymers show spectroscopic properties (NMR, IR, UV) in accordance with the proposed structures. A glass transition temperature (Tg) of 92°C and a thermal degradation temperature (Td) of 322°C were recorded. The poled polymer film exhibits stable second-order nonlinear optical activity (d33 = 4.2 pm/V) over a period of 800 hours as characterized by the temporal response of the second harmonic signal at room temperature.

Interfacing Conducting Polymers and Biological Macromolecules: A Case Study of Insecticide Biosensor Development

The creation of smart materials is a current goal of many laboratories including our own. We are involved in studying a number of thin film and monolayer systems which share the common features of involving highly conjugated conducting organic polymers and the evolved properties of specific biological macromolecules. In this report we describe a generic ‘cassette’ methodology for immobilizing biotinylated biological macromolecules to hydrophobic surfaces using a novel class of conducting copolymers of polythiophene. These copolymers are derivatized with long alkyl chains and biotin moieties to bind, respectively, to the hydrophobic surface and the biotinylated species, through the biotin - streptavidin interaction. We utilize the monolayer ‘cassette’ approach to attach a signal transducing biomolecule alkaline phosphatase to the surface of a glass capillary. This produces a flexible system in which chemiluminescence provides the basis for a useful measurement strategy. A novel technique involving the generation of chemiluminescence signal from alkaline phosphatase catalyzed dephosphorylation of a macrocyclic phosphate compound is described. Detection of paraoxon and methyl parathion, both enzyme inhibitors, has been achieved at ppb levels. The technique is rapid, sensitive, and is applicable to the detection of all organophosphorus based insecticides. The technique will be used in developing a fiber;optic biosensor for remote detection of insecticides.

Photoconductivity and Photovoltage Generation in Novel Photocrosslinkable Nonlinear Optical Polymers

Abstract : Photoconductivity in photocrosslinkable second order nonlinear optical (NLO) polymeric materials and photovoltage generation in a poled guest- host polymer are reported. These NLO polymers have been functionalized with crosslinkable cinnamoyl groups and show relatively stable second order optical nonlinearities at room temperature when poled and crosslinked. Without introducing any photosensitizer or charge carrier transport agents, photoconductivity was obtained in these polymer systems. Optical excitation of the NLO chromophores lead to photocarrier generation and it is conjectured that these NLO chromophores also play a role in the carrier transport. In addition to photoconductivity, photovoltage generation was observed in the dye doped polymer upon poling. This poled polymer also exhibits a polarization dependent photovoltage when illuminated by light in the absorption region of the NLO chromophore.... Photocrosslinkable nonlinear optical polymer, Pphotovoltage generation, Guest host.

Biotinylated Polythiophene Copolymer – a Novel Electroactive Biomaterial Utilizing the Biotin-Streptavidin Interaction

Abstract : A novel hierarchical biomaterial capable of incorporating any biotinylated biomolecule has been created. Our strategy is to biotinylate one- dimensional electroactive polymers and use a bridging streptavidin protein on Langmuir-Blodgett (LB) organized films. The following copolymeric system which enables functionalization of other molecules and formation of good monolayers was employed. Biotinvlated poly(3-methanolthiophene-co-3-undecylthiophene (B- PMUT) demonstrated a significantly better isotherm implying superior molecular packing compared to poly(3-methanol thiophene-co-3-undecyl thiophene) (PMUT) on the LB air-water surface. The isotherm showed significant area expansion when streptavidin was injected below the B-PMUT monolayer in 0.1 mM NaH2PO4/0-1 M NaCl buffer (ph 6.8) subphase. We then incorporated biotinylated phycoerythrin (B-PE) into this novel biomaterial by binding the unoccupied biotin binding sites on the bound streptavidin (4 sites total). The pressure-area isotherm of the protein injected monolayer showed area expansion. A characteristic fluorescent emission peak at 576 nm was detected from the monolayer transferred onto a solid substrate. These observations demonstrated the function of B-PMUT in hierarchical monolayer assembly of molecules incorporating the biotin/ streptavidin interaction. Langmuir-Blodgett, Electroactive polymers, Biotin, Streptavidin, Phycoerythrin.