Photoactivation: the assembly of the active site of the water oxidising enzyme

Completed

Renewable Energy Sources(Bio-Energy, Other bio-energy)

Basic and strategic applied research

BIOLOGICAL AND AGRICULTURAL SCIENCES (Biological Sciences)

Not Cross-cutting

Prof W Rutherford
Life Sciences - Biology
Imperial College London

Research Grant

BBSRC

01 October 2012

30 September 2015

36 months

£367,556

Transport Systems and Vehicles

London

Prof W Rutherford, Life Sciences - Biology, Imperial College London

The research will elucidate the process of photoactivation: the assembly of the water oxidising enzyme, Photosystem II (PSII). This occurs when the enzyme is made and when it is repaired. PSII is the main solar energy converter on Earth: it puts the energy into biosphere. It is made in vast quantities and each one is repaired every 30 minutes. Photoactivation limits plant growth and survival of the organism under some stress conditions. Understanding this process is thus important for agriculture for the production of food, fibre and fuel. The enzyme itself is the fastest of any water oxidation catalyst made from cheap, Earth-abundant materials and it thus represents the bench-mark model for catalyst design for solar fuel production in artificial photosynthesis. This is considered to be one of the very few scalable, sustainable technologies that will replace fossil fuels. The enzyme is responsible for putting the O2 into the atmosphere, thereby changing the nature of life and the planet itself. The events in photoactivation probably reflect the evolution of the enzyme, with early steps in the assembly of metal cluster representing evolutionary stages in the enzyme's development. Overall then the project is of intrinsic interest and benefit to a very wide public. In the energy sector, beneficiaries include: companies wishing to develop alternatives to fossil fuels and those who wish to move into green chemistry; governments and policy makers who wish for energy security and for new energy sources for developing countries; the armed forces who are looking for alternative fuels for specific and niche uses; environmentalists who need to focus on rational long-term alternatives to fossil fuels; and the general public who will soon have to accept that a change to a sustainable energy regime is inevitable. The benefits derived from the development of artificial photosynthesis as an alternative technology are evident and cannot be overestimated. They will contribute to mitigating the impending climate/energy crisis: the biggest problem facing mankind. If this problem is not solved, then all other problems of health, economy, etc will soon be dwarfed by it. Bioenergies will also contribute to the sustainable energy sector, although this is likely to be on a much smaller scale and mainly for niche uses. Bioenergies are mainly based on photosynthesis either directly or indirectly, so PSII research is central to them. The present project will be of benefit to bioenergy companies, farmers, policy makers, governments and the armed forces. In the agricultural sector, beneficiaries include: companies involved in developing new species, strains or processes for maintaining and or improving crop yields under stress conditions of changing environments; farmers who wish to develop new practices for similar reasons; governments and policy-makers interested achieving food security; and the public who will benefit from food security. In the environmental and ecologica l sector, beneficiaries include: those wishing to understand and mitigate loss of photosynthetic species in a changing environment. These include professionals in the sector, governments and policy makers, the tourism sector (coral reef bleaching) and fisheries (the loss of photosynthetic microbes at the start of the food chain). In the education sector, in museums and in the media etc, there will be benefits from the intrinsic interest in one of the few definable milestones in the origins and evolution of life: the ability to oxidise water resulting in the appearance of O2 in the environment. Staff hired for the project will obtain training in cutting-edge research in a leading lab in this important area. They will have the benefit of the excellent intellectual environment of a leading university with the tradition of close ties with engineers and applications. It is likely that they will go on to contribute to this subject in UK industry or academia.

Objective 1. Photoactivation of PSII in the thermophilic cyanobacterial PSII: establishing the optimal physical, chemical and biochemical conditions. The flash-pairs method of Chenaie will be used with bacterial cells and isolated enzyme to obtain the two key kinetic parameters for the initial steps of photoactivation. We shall also determine the temperature and O2 dependencies of these processes. We shall test the influence of extrinsic polypeptides on photoactivation.
Objective 2. Generation and characterisation of mononuclear Mn intermediates in PSII photoactivation in T. elongatus: the first steps. The first Mn3+ state will be generated by flash and cw illumination under a range of conditions. Various components (Mn2+, Mn3+, Mn4+, TyrZ, TyrD and the electron acceptors) will be monitored by different types of EPR, kinetic UV/visible and fluorescence spectroscopies before and after the B to C step.
Objective 3. Generation of multinuclear Mn intermediates in photoassembly: the hunt for S-4, S-2 etc. We shall use EPR to search for multinuclear Mn complexes formed as intermediates during photoactivation. We will focus on Mn3+Mn2+ dimer states including the S-2 state. Understanding these states could allow us to distinguish between the high valence and low valence models for the functional Mn. The effect of Ca2+ binding will be studied before and after formation of the high valence intermediates.
Objective 4 Structural characterisation of intermediates in photoactivation. When intermediates are generated that are sufficiently stable, PSII shall be the subject of crystallisation trials. The Mn-depleted enzyme, monomeric and dimeric forms, will be studied with and without assembly proteins, e.g. Psb27, PsbP and PsbQ. Mn-free PSII is a target as a basis for understanding the changes occurring during photoactivation.

14/04/14