Nanofabricated media with negative permeability at visible frequencies

A great deal of attention has recently been focused on a new class of smart materials-so-called left-handed media-that exhibit highly unusual electromagnetic properties and promise new device applications. Left-handed materials require negative permeability ν, an extreme condition that has so far been achieved only for frequencies in the microwave to terahertz range. Extension of the approach described in ref. 7 to achieve the necessary high-frequency magnetic response in visible optics presents a formidable challenge, as no material-natural or artificial-is known to exhibit any magnetism at these frequencies. Here we report a nanofabricated medium consisting of electromagnetically coupled pairs of gold dots with geometry carefully designed at a 10-nm level. The medium exhibits a strong magnetic response at visible-light frequencies, including a band with negative ν. The magnetism arises owing to the excitation of an antisymmetric plasmon resonance. The high-frequency permeability qualitatively reveals itself via optical impedance matching. Our results demonstrate the feasibility of engineering magnetism at visible frequencies and pave the way towards magnetic and left-handed components for visible optics. © 2005 Nature Publishing Group.

LacO-LacI interaction in affinity adsorption of plasmid DNA

Current approaches for purifying plasmids from bacterial production systems exploit the physiochemical properties of nucleic acids in non-specific capture systems. In this study, an affinity system for plasmid DNA (pDNA) purification has been developed utilizing the interaction between the lac operon (lacO) sequence contained in the pDNA and a 64mer synthetic peptide representing the DNA-binding domain of the lac repressor protein, LacI. Two plasmids were evaluated, the native pUC19 and pUC19 with dual lacO3/lacOs operators (pUC19lacO3/lacOs), where the lacOs operator is perfectly symmetrical. The DNA-protein affinity interaction was evaluated by surface plasmon resonance using a Biacore system. The affinity capture of DNA in a chromatography system was evaluated using LacI peptide that had been immobilized to Streamline™ adsorbent. The KD-values for double stranded DNA (dsDNA) fragments containing lacO1 and lacO3 and lacOs and lacO3 were 5.7 ± 0.3 × 10 -11 M and 4.1 ± 0.2 × 10-11 M respectively, which compare favorably with literature reports of 5 × 10-10 - 1 × 10-9 M for native laCO1 and 1-1.2 × 10-10 M for lacO1 in a saline buffer. Densitometric analysis of the gel bands from the affinity chromatography run clearly showed a significant preference for capture of the supercoiled fraction from the feed pDNA sample. The results indicate the feasibility of the affinity approach for pDNA capture and purification using native protein-DNA interaction. © 2006 Wiley Periodicals, Inc.

Investigation of the activation mechanism and structural characterization of the CGRP receptor

The calcitonin gene-related peptide (CGRP) receptor is an unusual G protein-coupled receptor (GPCR) in that it comprises the calcitonin receptor-like receptor (CLR), receptor activity modifying protein 1 (RAMP1) and the receptor component protein (RCP). The RAMP1 has two other homologues – RAMP2 and RAMP3. The endogenous ligand for this receptor is CGRP, a 37 amino acid neuropeptide that act as a vasodilator. This peptide has been implicated in the aetiology of health conditions such as inflammation, Reynaud’s disease and migraine. A clear understanding of the mode of activation of this receptor could be key in developing therapeutic agents for associated health conditions. Although the crystal structure of the N-terminal extracellular domain (ECD) of this receptor (in complex with an antagonist) has been published, the details of receptor-agonist interactions at this domain, and so ultimately the mechanism of receptor activation, are still unclear. Also, the C-terminus of the CLR (in the CGRP receptor), especially around the presumed helix 8 (H8) region, has not been well studied for its role in receptor signalling. This research project investigated these questions. In this study, certain residues making up the putative N-terminal ligand-binding core of the CLR (in the CGRP receptor) were mapped out and found to be crucial for receptor signalling. They included W69 and D70 of the WDG motif in family B GPCRs, as well as Y91, F92, D94 and F95 in loop 2 of CLR N-terminus. Also, F163 at the cytoplasmic end of TM1 and certain residues spanning H8 and associated C-terminal region of CLR were found to be required for CGRP receptor signalling. These residues were investigated by site-directed mutagenesis where they were mutated to alanine (or other residues in specific cases) and the effect of the mutations on receptor pharmacology assessed by evaluating cAMP production, cell surface expression, total cell expression and aCGRP-mediated receptor internalization. Moreover, the N-terminal ECDs of the CLR and RAMPs (RAMP1, RAMP2 and RAMP3) were produced in a yeast host strain (Pichia pastoris) for the purpose of structural interaction study by surface plasmon resonance (SPR). Following expression and purification, these receptor proteins were found to individually retain their secondary structures when analysed by circular dichroism (CD). Results were analysed and interpreted with the knowledge of the secretin family receptor paradigm. The research described in this thesis has produced novel data that contributes to a clearer understanding of CGRP receptor pharmacology. The study on CLR and RAMPs ECDs could be a useful tool in determining novel interacting GPCR partners of RAMPs.

Biosensor based on excessively tilted fiber grating in thin-cladding optical fiber for sensitive and selective detection of low glucose concentration

We report a highly sensitive, high Q-factor, label free and selective glucose sensor by using excessively tilted fiber grating (Ex-TFG) inscribed in the thin-cladding optical fiber (TCOF). Glucose oxidase (GOD) was covalently immobilized on optical fiber surface and the effectiveness of GOD immobilization was investigated by the fluorescence microscopy and highly accurate spectral interrogation method. In contrast to the long period grating (LPG) and optical fiber (OF) surface Plasmon resonance (SPR) based glucose sensors, the Ex-TFG configuration has merits of nearly independent cross sensitivity of the environmental temperature, simple fabrication method (no noble metal deposition or cladding etching) and high detection accuracy (or Q-factor). Our experimental results have shown that Ex-TFG in TCOF based sensor has a reliable and fast detection for the glucose concentration as low as 0.1~2.5mg/ml and a high sensitivity of ~1.514nm·(mg/ml)−1, which the detection accuracy is ~0.2857nm−1 at pH 5.2, and the limit of detection (LOD) is 0.013~0.02mg/ml at the pH range of 5.2~7.4 by using an optical spectrum analyzer with a resolution of 0.02nm.