Use this URL to cite or link to this record in EThOS: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.495327
Title: Protein refolding via immobilisation on crystal surfaces
Author: Davidson, Katrina Ann
Awarding Body: University of Glasgow
Current Institution: University of Glasgow
Date of Award: 2008
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Abstract:
Is it possible to find an easy, generic method for protein refolding? The preparation of functionally active protein molecules from the unfolded state can be a difficult task. Although there are many well-established techniques for protein refolding, such as dilution, dialysis, chromatography and others, in many instances these methods can be time consuming and inefficient. A rapid, inexpensive and simple method for protein folding is a much sought after technique. Proteins in the unfolded state (either inclusion bodies or unfolded by chemical or physical means) are generally solubilised in solutions containing urea or guanidine hydrochloride. The removal of these molecules from the protein environment is commonly utilised as a method for triggering refolding. A new method for the refolding of biomolecular species has been developed via the formation of Protein Coated Micro-crystals (PCMC). The formation of PCMC is a recently developed method for the immobilisation protein upon the surface of a watersoluble excipient (salt, amino acid or sugar) via a co-precipitation reaction in a water miscible organic solvent. These proteins can then be used as immobilised biocatalysts in both the aqueous and organic phase. In the immobilisation of unfolded, solubilised protein, the solubilising agents (e.g. urea or guanidine hydrochloride) are removed from the protein environment as they are soluble in the organic phase. The removal of these molecules initiates protein folding during the coprecipitation process. In the course of this project, a number of proteins were studied in order to observe their behaviour in this immobilisation and simultaneous folding process. Lysozyme was utilised as it is an enzyme which is relatively simple to refold from the chemically unfolded state by conventional methods such as dilution. Upon immobilisation of lysozyme from the chemically unfolded state, up to 92% of the activity of the native protein was regained. The enzyme lipase, which is notoriously difficult to fold, was also used to determine the efficiency of this method under more challenging conditions. Lipase immobilised from the chemically unfolded state was seen to regain up to 36 % of the activity of the native protein.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.495327  DOI: Not available
Keywords: QD Chemistry
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