Dr Alethea Tabor
The common theme of the projects in the Tabor group is the application of organic chemistry to the solution of biological problems. We synthesise both naturally occurring biologically important molecules, and design and synthesise non-natural and labelled analogues which can be used as tools to solve structural and mechanistic problems in biological chemistry, and can also be used to probe biological pathways in vitro and in vivo . These projects are synthetically challenging and, combined with biophysical and molecular biological techniques provide us, and our biological collaborators, with a range of tools not normally available for the study of biological systems.
Total synthesis and biological activity of the lantibiotics
The lantibiotics are an important class of antibiotic peptides with complex bridged structures formed by a series of thioether links between the amino acid side chains. An example is nisin .
In spite of the pressing need for new classes of antibiotics for use against resistant bacteria, these have been little explored because of the difficulties inherent in their synthesis. We have developed a new, direct route to the parent amino acid, lanthionine, which avoids problems encountered in previous approaches. We have then developed a new method for synthesising lantibiotics using solid-phase peptide synthesis techniques
We are using this approach to prepare peptides with multiple thioether bridges, and even with overlapping thioether bridges. We are currently applying this to the synthesis of subunits of nisin, and investigating how the sequence of each subunit of nisin affects the conformation of the ring and the binding of nisin to the biological targets.
Conformationally restricted peptides of defined secondary structure
We have recently developed a powerful general method for the solid-phase synthesis of peptides bearing unnatural linkages between two C a - positions. This method allows the on-resin synthesis of polycyclic peptides, which were not previously accessible, and also allows a much wider range of conformational constraints to be studied. We have so far synthesised peptides with an aliphatic linkage between the i and i+4 positions. These were designed as a -helix mimics: in contrast to the literature, however, they adopt type I b -turn conformations. We are currently synthesising peptide with different linkages in order to explore and exploit this effect .
Non-viral gene therapy using a cell-targeted ternary vector.
In a major interdisciplinary research project, we are elucidating the structure and mode of action of a novel, highly potent non-viral gene delivery system, and designing new non-viral gene delivery vectors based on this system. The work is being carried out in collaboration with Dr S. L Hart (Institute of Child Health, UCL), Dr H C Hailes (Chemistry), Professor M J Lawrence (School of Pharmacy, KCL), and Dr D Zicha (ICRF).
These targeted, self-assembling lipid:peptide (lipopolyplex) vector complexes (LID) consist of a lipid component, Lipofectin (L) (the cationic lipid DOTMA with a co-lipid DOPE in a 1:1 ratio), plasmid DNA (D), and a dual-function, cationic peptide component (I) comprising both a K 16 (DNA condensation) and a cell-targeting sequence.
LID systems display high transfection efficiency and low toxicity in vitro and in vivo , transfect non-dividing cells efficiently, and are well tolerated with low immunogenicity in vivo . Our working hypothesis for the transfection pathway is that the LID complexes bind to specific integrins on the cell surface and are internalised by receptor-mediated endocytosis. The lipid component then mediates endosomal fusion, releasing the peptide/DNA complex into the cytoplasm: the peptide then mediates transport into the nucleus, followed by transcription.
Using fluorescently labelled vector components, and biophysical techniques (FCS, FRET, fluorescence quenching and cryo-EM), we have worked out the macromolecular organisation of the three components (peptide, lipid and DNA) within the vector.
This is the first time that a nanostructure of this complexity has been analysed in this detail.
We are synthesising new peptide, lipid and lipopeptide structures in order to improve the efficiency and stability of this vector. The rationale behind much of the design is to understand and imitate those features of viruses that make them such efficient gene transfer agents - e.g. superior serum stability, cell-specific targeting and receptor-mediated endocytosis, enhanced transport of the internalised DNA to the nucleus.
For each enhancement of each component, we are carrying out systematic studies of the biophysics, intracellular transport and cell-specific transfection properties of the resulting complex in order to understand and precisely define the effects that structural changes to each component has on the activity of the vector.
Delivery of Enzyme Inhibitors to Dendritic Cells
We are working with Professor B. Chain (Immunology) on a novel approach to delivery of enzyme inhibitors via receptor-mediated endocytosis. We have so far successfully used mannose-pepstatin conjugates with cleavable linkers to specifically delivery this highly insoluble inhibitor to dendritic cells, and have specifically knocked out the action of cathepsin E (in cat D-deficient mice). We have recently been awarded a BBSRC grant (SCIBS Initiative) to extend this work to the examination of localisation of the enzyme and inhibitor, and to use other inhibitors to examine other pathways of antigen processing and presentation.
Cell culture systems
We are working on the development of scalable biphasic co-culture systems with Dr Farlan Veraitch (Biochemical Engineering, UCL). This project involves designing and synthesising biocompatible materials modified with ligands that will promote, or inhibit, cellular differentiation and proliferation.
In collaboration with Dr Daren Caruana we are designing polymeric amino acid-based materials, with defined secondary structure and bearing electrochemical reporter groups, as electrochemical sensors.
- B. M. Chain, P. F. Free, P. Medd, C. Swetman, A. B. Tabor, N. Terrazzini, 'The expression and function of cathepsin E in dendritic cells'
J. Immunol ., 174 , 1791-1800 (2005).
- S. Bregant, A. B. Tabor, 'Orthogonally Protected Lanthionines: Synthesis and Use for the Solid-Phase Synthesis of an Analogue of Nisin Ring C'
J. Org. Chem . 70 , 2430 -2438 (2005).
- S. Sarkar, L. K. Lee, S. L. Hart, H. C. Hailes, S. M. Levy, A. B. Tabor , P. Ayazi Shamlou, 'Fractal Structure of Polycation-DNA Complexes'
Biotech. Appl. Biochem. 41 , 127-136 (2005).
- J. F. Markusen, C. Mason, D. A. Hull, M. A. Town, A. B. Tabor, M. Clements, C. Boshoff and P. Dunnill, 'The Behaviour of Adult Human Mesenchymal Stem Cells Entrapped in Alginate-GRGDY Beads' Tissue Engineering, 12, 821-830 (2006).
- P. F. Free, C. A. Hurley, T. Kageyama, B. M. Chain, A. B. Tabor, 'Mannose-Pepstatin Conjugates as Targeted Inhibitors of Antigen Processing' Org. Biomol. Chem., 4, 1817-1830 (2006).
- I. Tranchant, A.-C. Herve, S. Carlisle, P. Lowe, C. J. Slevin, C. Forssten, J. Dilleen, D. E. Williams, A. B. Tabor, H. C. Hailes, 'Design and synthesis of ferrocene probe molecules for detection by electrochemical methods'
Bioconj. Chem., 17, 1256 - 1264 (2006).
- M. A. Pilkington-Miksa, M. J. Writer, S. Sarkar, Q.-H. Meng, S. E. Barker, P. Ayazi Shamlou, S. L. Hart, H. C. Hailes, A. B. Tabor, 'Targeting Lipopolyplexes Using Bifunctional Peptides Incorporating Hydrophobic Spacer Amino Acids: Synthesis, Transfection and Biophysical Studies' Bioconj. Chem., 18, 1800 - 1810 (2007).
- M. F. Mohd Mustapa, P. C. Bell, C. A. Hurley, A. Nicol, E. Guénin, S. Sarkar, M. J. Writer, S. E. Barker, J. B. Wong, M. A. Pilkington-Miksa, B. Papahadjopoulos-Sternberg, P. Ayazi Shamlou, H. C. Hailes, S. L. Hart, D. Zicha, A B. Tabor, 'Biophysical characterization of an integrin-targeted lipopolyplex gene delivery vector'
Biochemistry, 46, 12930 12944 (2007).
- M. A. Pilkington-Miksa, S. Sarkar, M. J. Writer, S. E. Barker, P. Ayazi Shamlou, S. L. Hart, H. C. Hailes, A. B. Tabor, 'Synthesis of Bifunctional Integrin-Binding Peptides Containing PEG Spacers of Defined Length for Non-Viral Gene Delivery' Eur. J. Org. Chem, 2900 - 2914 (2008).
- M. F. Mohd Mustapa, S. M. Grosse, L. Kudsiova, M. Elbs, E.-A. Raiber, J. B.Wong, A. P. R. Brain, H. E. J. Armer, A. Warley, M. Keppler, T. Ng, M. J.Lawrence, S. L. Hart, H. C. Hailes, A. B. Tabor, 'Stabilized Integrin-Targeting Ternary LPD (Lipopolyplex) Vectors for Gene Delivery Designed To Disassemble Within the Target Cell'Bioconj.Chem., 20, 518 532 (2009).