In nature, a number of organisms and organelles are capable of self-propulsion at the micro- and nano-scale. Inspired by biological motors, this investigation aims to induce self-propulsion of an enzyme, by the biocatalytic self-assembly of aromatic peptide amphiphile molecules into supramolecular fibre structures. The individual motion of enzymes is measured directly using fluorescence microscopy, by the covalent attachment of alkaline phosphatase to fluorescing quantum dots. Enzyme-quantum dot conjugates represent nanoparticulate ‘vehicles’ transported by the enzyme 'motor'. Two aromatic peptide substrate ‘fuels’, initially assembled in a micellar form, are studied for their ability to propel enzyme-quantum dot conjugates, by biocatalytic dephosphorylation causing self-assembly into one-dimensional fibres. The effect of the ‘fuel’ on conjugate motion is compared with controls consisting of no fuel, a non-self-assembling substrate and a non-directional self-assembling substrate (i.e. one that assembles into spheres, but not fibres). Significant quantities of data were obtained for each substrate scenario and speed distribution plots revealed that enzyme-conjugates exhibit faster transport with the fibre forming system, compared to controls. Further to this, upon increasing the concentration of the fibre-forming fuel, the average speed of the conjugates increases, although directionality remains random. An initial investigation for directional control is carried out using 'fuel' reservoirs consisting of substrate saturated polyacrylamide gels. Substrates diffuse from the gel into surrounding motility medium creating a concentration gradient, which the enzyme-motors are proposed to travel along in a directional manner. The proposed propulsion model for self-assembly-driven motion of enzyme-conjugates is that short bursts of fibre growth provides linear propulsion which increases the diffusion rate of the enzyme-conjugate. Simultaneous visualisation of self-assembled fibres and enzyme-quantum dot conjugates is attempted, using extrinsic and intrinsic fluorescent methods, to investigate the mechanism proposed for fibre-propulsion. Finally, enzymes thermolysin and α-chymotrypsin are investigated as a step toward generalising the method for other enzymes and for their potential use in a multi-enzyme/multi-coloured quantum dot system for future applications in nano-separation of enzymes.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:665239 |
| Date | January 2015 |
| Creators | Leckie, Joy Susan |
| Publisher | University of Strathclyde |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Source | http://oleg.lib.strath.ac.uk:80/R/?func=dbin-jump-full&object_id=25502 |
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