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Title: Molecular level investigations of coiled-coil proteins
Author: Johnson, Daniel
ISNI:       0000 0001 3591 082X
Awarding Body: University of Nottingham
Current Institution: University of Nottingham
Date of Award: 2006
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The coiled-coil is a very common protein structural motif, consisting of two or more alpha helices intertwined in a supercoil. In biological systems it is found in proteins which fulfil a number of roles, for example in structural proteins such as keratin, as well as a large number of transcription factors and other DNA binding proteins, where it functions as a dimerisation domain. This versatile motif has also been adapted for a number of applications including the production of responsive hydrogels; the construction of self-assembling fibres for tissue engineering applications; as a cross-linking agent in drug delivery applications, as well as in the creation of biosensor surfaces. In this study two different types of protein containing coiled-coil domains are examined. It is the aim of this work to increase the understanding of how behaviour of this motif at the molecular level relates to those at the macroscopic. The first set of proteins of interest in this thesis form pH-responsive hydrogel systems utilising a variant of the coiled-coil motif, the leucine zipper, as their cross-linking domain. These consist of two or more leucine zippers separated by random coil spacer sections. When the leucine zipper sequences dimerise, they cross-link the proteins forming a gel matrix. As charges within the leucine zippers alter, as the pH of their environment is altered, their relative stability changes causing them to associate or dissociate. This is reflected in a change in the physical characteristics of gels from visco-elastic solids to viscous liquids which is reversible. The bulk properties of these materials is well characterised, but relatively little information is known at the molecular level. Starting at the single molecule level and scaling up to the mesoscale, the behaviour of hydrogel-forming proteins has been probed using a range of techniques, particularly with respect to the interactions of the coiled-coil forming domains. Studies described in this thesis have concentrated on the change of physical characteristics of these proteins and their assemblies as pH was altered. We have thus been able to connect behaviour of these proteins at the molecular scale to bulk properties of solutions and materials formed from them. In the final experimental chapter a coiled-coil structure of more complex architecture is examined. This is formed of a ternary complex of three proteins which assemble into a two-stranded leucine zipper domain, designed as a cross-linking motif for nano-particle assemble. Force was used to probe the kinetic stability and mechanical strength of this assembly. With the increasing use of the coiled-coil motif as a cross-linking domain in the use of biomaterials it is increasingly important to be able to connect the behaviour of such materials to the behaviour of the coiled-coil motif at the molecular scale, to gain further insight into the mechanistics behind their behaviour.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available
Keywords: QP501 Animal biochemistry