Use this URL to cite or link to this record in EThOS:
Title: Structure-function analysis of the plant alternative oxidase expressed in Schizosaccharomyces pombe mitochondria
Author: Crichton, P. G.
ISNI:       0000 0004 2718 089X
Awarding Body: University of Sussex
Current Institution: University of Sussex
Date of Award: 2004
Availability of Full Text:
Access from EThOS:
The alternative oxidase (AOX) is a terminal respiratory oxidase found in the mitochondria ()f higher plants, many fungi as well as several protists. Its activity is non-protonmotive and therefore decreases the efficiency of mitochondrial energy transduction. Structural and mechanistic insights into AOX regulation and catalysis are therefore important for a I'undamental understanding of mitochondrial energy metabolism. Given that some parasites. unlike their mammalian hosts, are dependent upon an AOX, such information may also facilitate the design of novel AOX inhibitors as potential chemotherapeutic drugs. The work presented in this thesis comprises a biochemical analysis of plant AOX proteins expressed in isolated yeast mitochondria using a previously established heterologous system. An initial characterisation of isolated S. pombe mitochondria reveals oxygen uptake activity dependent upon the presence of ethanol. This activity is shown to be the result of a novel matrix-located alcohol dehydrogenase that regenerates NADH, which is subsequently oxidised by the respiratory chain. Data obtained using a system kinetic approach provide further insight into the unusual behaviour of this respiratory pathway. The relative activities of site-specific mutants of the Sauromatum guttatum AOX have been assessed in isolated yeast mitochondria and characterised with respect to substrate and inhibitor affinity. The mutation of two residues is shown to affect the apparent affinity of AOX for oxygen. A potential influence of these residues on the active site of AOX is discussed. Following the expression of a second AOX isozyme (AOXla from Arabidopsis thaliana), respiratory analysis with isolated yeast mitochondria and mitochondrial membranes indicates that the S. guttatum AOX is the first plant isozyme identified that is not regulated by organic acids and appears to be in a constitutively active state. A bioinformatic study implicates novel residues, other than established regulatory cysteines, in the interaction with organic acids and provides a framework for structural hypotheses
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available