Peptide aptamers are alternative binding proteins, whose development is motivated by the desire to overcome the drawbacks of antibodies. Defined as variable peptides constrained by a constant protein scaffold, peptide aptamers ., have been developed by several groups over the past two decades and this thesis will detail my and my colleagues' contribution to the field. Chapter 1 will provide the reader with a brief account of the history and applications of peptide aptamers, as well as give a rational for the technology. In the second chapter the development of SQM, a novel peptide aptamer scaffold derived from the protease inhibitor Stefin A, is described. SQM is based on the previously engineered STM scaffold, but, different to its predecessor, is capable of presenting randomised peptides from three different insertion sites. The stability and functionality of the new scaffold is explored, a process which highlights some structural problems in SQM. In a successful attempt to overcome these issues SQM is re engineered to yield SQT and several applications of the new scaffold, such as the generation of large combinatorial peptide aptamer libraries, are described (chapter 3). In order to further the peptide aptamer field the advancement of auxiliary technologies, such as affinity peptide tags for protein detection in western blots, immunocytochemistry etc., is crucial. Thus, in chapter 4, the development of an improved streptavidin-binding tag - generated by simply exploiting the multimeric nature of streptavidin - is outlined. In the final chapter I make use of the biologically neutral SQT scaffold to present and constrain four different apoptotic BH3 domains, with the aim of generating new tools for the study of the intrinsic apoptotic pathway. In combination the work presented here establishes Stefin A derived peptide aptamers as viable binding proteins with potential applications in a range of laboratory and clinical settings.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:550354 |
| Date | January 2012 |
| Creators | Stadler, Lukas Kurt Josef |
| Publisher | University of Leeds |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
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