Nanodissection of protein-protein interactions in energy transducing membrane protein complexes

Lead Research Organisation: University of Sheffield
Department Name: Molecular Biology and Biotechnology

Abstract

The major energy transducing reactions in the biosphere are mediated by membrane protein complexes that convert and store solar energy as a protonmotive force (photosynthesis), or sequentially transfer electrons to oxygen (respiration). Each of these globally important processes requires that a membrane-extrinsic electron carrier docks onto a large, membrane -bound complex, as for example with cytochrome c and cytochrome aa3 oxidase in mitochondria. Although the structures of all of the photosynthetic or respiratory components are known, little is known of the intermolecular forces involved in establishing transient electron transfer complexes at the membrane surface. Moreover, the protein-protein and protein-cofactor interactions that stabilize these multisubunit complexes are also unknown.

Publications

10 25 50

Studentship Projects

Project Reference Relationship Related To Start End Student Name
BB/M011151/1 01/10/2015 30/09/2023
1688092 Studentship BB/M011151/1 01/10/2015 30/09/2019 David Farmer
 
Description The work reported in this thesis has used a range of biochemical, biophysical and structural tech-
niques to study these biologically important and mechanistically unique enzymes: microscale ther-
mophoresis, kinetic analysis and cross-linking mass spectrometry revealed that the driving force
behind chelation, the ATPase activity of the AAA+ ChlI subunit, is linked to the chelating ChlH
protein via the regulatory ChlD subunit; X-ray crystallography-guided mutagenesis determined
the porphyrin binding site in ChlH; (cryo)-electron microscopy was used to initiate structural in-
vestigations into the quaternary organisation of MgCH and POR; and 2-dimensional electronic
spectroscopy was applied to the light-initiated reaction catalysed by POR, the first use of this com-
plex technique on an enzyme, potentially revealing a novel intermediate in the reaction that is
formed on the femtosecond time-scale. The work presented in this thesis aims to develop our bio-
chemical understanding of MgCH and POR, and lay the foundations for further structural and
mechanistic studies of these interesting enzymes of chlorophyll biosynthesis.
Exploitation Route It has laid the ground work for further structural studies by cryo-EM and many people in the lab are able to do cryo-EM now due to training from me.
Sectors Agriculture, Food and Drink,Chemicals,Energy,Environment,Manufacturing, including Industrial Biotechology