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Engineering biological magnetic particles for biotechnology

Principal Supervisor: Dr Tim Overton, School of Chemical Engineering

Co-supervisor: Owen Thomas, School of Chemical Engineering

PhD project title: Engineering biological magnetic particles for biotechnology

University of Registration: Birmingham

Project outline:

Magnetotactic bacteria (MTB) were first identified over 50 years ago [1]. This class of bacteria contains nanoscale membrane-bound magnetic particles called magnetosomes. Magnetosomes are naturally-occurring nanoscale organelles that allow magnetotactic bacteria to physically align themselves with the earth’s magnetic field, and comprise an iron-containing magnetic core (Fe3O4 or Fe3S4) surrounded by a membrane studded with proteins. The structure of magnetosomes allows engineering of functionality for a range of biotechnological applications. It is possible to engineer the magnetosome surface proteins to have desirable functionality, for example by fusion with green fluorescent protein or ligands directed towards a desired target such as biotin-streptavidin.

The past decade has seen explosive growth in the use of magnetic particle-based techniques for the capture and purification of huge variety of biological entities (from molecules to cells), and the recovery and recycling of biological catalysts, e.g. immobilised enzymes, cells or parts of cells [2,3]. Central to both types of application is the provision of suitable magnetic support materials [2-4]. To date these are produced by chemical means in multiple, often complex and costly steps [2-6].

Magnetosomes offer the promise of a biologically-synthesised magnetic particle with customisable functionality, generated in a cleaner, greener, and potentially cheaper manner than that currently employed.

In this project: the student will investigate the engineering and use of magnetosomes for biotechnological purposes, with a focus on the generation of magnetic particles for the capture and purification of biopharmaceutical drug products such as antibody fragments.

‘State-of-the art’ knowledge on the molecular biology [7] and cultivation of magnetotactic bacteria in fermenters [8,9] will be employed to create new classes of magnetic particles potentially ideal for a wide range of biotech applications (e.g. preparative separation of high value proteins, magnetic affinity cell sorting, high-throughput diagnostic screening).

Several different classes of magnetosomes will be generated, including (from simple to more complex and challenging):

  1. base magnetosomes amenable to chemical derivatization with affinity ligands;
  2. biotin-binding magnetosomes for linkage to biotinylated ligands; and
  3. fully finished magnetic affinity adsorbents composed of magnetosomes displaying surface tethered affinity ligands.

The student shall:

  1. employ molecular biology based synthetic biology and fermentation tools for magnetotactic bacterial strain improvement and generation of recombinant magnetosomes displaying specific proteins (e.g. affinity, GFP) on their surfaces;
  2. optimise and
  3. intensify fermentative production of the above magnetosomes; and
  4. demonstrate effective use of magnetosome affinity adsorbents for the recovery and purification of selected target molecules from a variety of crude bioprocess feedstocks.

References: 

  1. Yan L et al. (2012) Microbiol Res 167: 507-519
  2. Franzreb M et al. (2006) Appl Microb Biotechnol 70: 505-516
  3. Peuker U et al. (2010) In Wiley Encyclopedia of Industrial Biotechnology, Bioprocess, Bioseparation and Cell Technology, ed. M.C. Flickinger, Wiley-Blackwell, Oxford, UK
  4. Fischer I et al. (2013) J Chromatogr A 1305: 7-16
  5. Franzreb M et al. (2006) In Process Scale Bioseparations for the Biopharmaceutical Industry, eds AA Shukla, MR Etzel, S Gadam, Taylor and Francis – CRC press
  6. Brown GN et al. (2013) Biotechnol Bioeng 110: 1714 –1725
  7. Pollithy A et al. (2011) Appl Environ Microbiol 77: 6165-6171
  8. Liu Y et al. (2010) Microb Cell Fact 9: 99 (doi: 10.1186/1475-2859-9-99)
  9. Zhang Y et al. (2011) Appl Environ Microbiol 77: 5851-5856

BBSRC Strategic Research Priority: Industrial Biotechnology and Bioenergy

Techniques that will be undertaken during the project:

The student will be trained in a range of skills encompassing diverse, but complementary fields to ensure that the project outcomes can be delivered, including:

  • Molecular biology – PCR, cloning, electrophoresis, DNA sequencing
  • Fermentation – use of bioreactors, optimisation and intensification of fermentation;
  • Downstream Processing – cell disruption, cross flow filtration (MF/UF/DF), adsorptive separation techniques such as high-gradient magnetic fishing and/or continuous magnetic extraction
  • Analysis methods - Electron and confocal laser scanning microscopy; FT-IR; HPLC; Gas MS; Flow cytometry; Gel electrophoresis; fluorescence, uv/vis and magneto spectrophotometry; vibrating sample magnetometry
  • Computational – Potential to use Design of Experiment and Surface Response approaches in R

Contact: Dr Tim Overton, University of Birmingham