Molecular Membrane Engineering for Nanoreactors

Membrane bound compartments are at the heart of biological complexity, allowing nature to control and drive biochemical reactions. The aim of this project is to create artificial bionanoreactors and linked vesicle networks that mimic this natural compartmentation. The approach is to engineer synthetic energy-transducing biomembrane vesicles that incorporate engineered membrane proteins and soluble components and to use light to generate electrochemical gradients to power and gate transport of materials between the nanoreactor and its environment. The ultimate goal is to control the flux of specific molecules into or out of the nanoreactor and the chemistry within them, enabling catalytic control for synthesis, degradation, delivery, sensation etc.

A key aspect of this work is to gain control over the composition of individual nanoreactors by developing component membrane proteins that self-assemble into hetero-oligomeric complexes with defined stoichiometry and orientation. A range of photosynthetic reaction centre membrane proteins modified with extra-membrane components such as coiled-coil bundles, the SpyCatcher/SpyTag system and the maltose binding protein have been engineered and are currently being assessed for expression, stability and their ability to form high affinity interactions. Their impact on reconstitution and sidedness in assembled proteoliposomes is also being studied through the development of a simple, convenient and generic assays based on the His-tags used to aid the purification of the various integral membrane proteins being used in the project. 

We are also developing the use of digital holographic microscopy, which measures refractive index changes, as a non-invasive measurement of transport. When coupled with conventional spectroscopic techniques, this will allow us to engineer nanoreactors with optimised influx and efflux of reactants and products in response to chemical and optical stimuli. We have established that the technique can be used to image individual liposomes of 1000 nm diameter and currently exploring methodologies for immobilising liposomes on surfaces for the application of measurements on single liposomes.