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Title: Prediction, design and characterisation of metal binding sites in antibodies
Author: Gregory, David St John
ISNI:       0000 0001 3518 1284
Awarding Body: University of Oxford
Current Institution: University of Oxford
Date of Award: 1992
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The design and creation of a functional metalloanti body is presented. A survey of metalloprotein structures was carried out to determine the common structural features of metal binding sites. Metal ligands were then introduced into hypervariable loop 11 of the anti-lysozyme antibody HyHEL-5 and the site comprehensively modelled. The final model was used as a basis for site-directed mutagenesis and mutant antibodies were produced using a Xenopus laevis oocyte expression system. Antigen binding studies showed the mutants to have an affinity for lysozyme equivalent to the parent antibody, while subsequent metal binding experiments have shown the new site to be capable of binding the transition metals cobalt, nickel, copper, zinc and cadmium. A method for the rapid prediction and design of metal binding sites in proteins is described. A protein structure is screened for the location of putative metal sites by template matching. After the introduction of suitable liganding residues, the metal binding potential at each site is evaluated using hydrophobicity contrast. A point representing the metal ion within a new site may be optimised with-respect-to the ligands using multidimensional minimisation. The method has been tested on known metalloprotein structures. It was routinely able to identify the metal binding sites and position a metal point in the site to within 0.6A of the actual metal ion.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available
Keywords: Metalloprotein structures