Award details

Probing the structure and function of a super-rogue photosystem II complex involved in chlorophyll f synthesis

Reference	BB/V002007/1
Principal Investigator / Supervisor	Professor Peter Nixon
Co-Investigators / Co-Supervisors	Dr James Murray
Institution	Imperial College London
Department	Life Sciences
Funding type	Research
Value (£)	594,088
Status	Current
Type	Research Grant
Start date	01/06/2021
End date	30/09/2024
Duration	40 months

Abstract

Synthesis of chlorophyll f (Chl f) in cyanobacteria requires the expression of the ChlF subunit, a paralogue of the D1 subunit of oxygen-evolving photosystem II, but the mechanism remains unclear. In background work we have discovered that ChlF is able to substitute for D1 to form a modified PSII complex with a role in Chl f synthesis rather than water oxidation. We have named this complex the super-rogue PSII complex (or sr-PSII). We have also identified a QD sequence motif in ChlF that is important for Chl f synthesis and possibly the binding of Chl f. To clarify the role of sr-PSII in Chl f synthesis, we propose to: (1) determine the co-factor composition of the sr-PSII complex; (2) use time-resolved absorbance and fluorescence spectroscopies to characterize its photochemical activity; (3) probe the presence of the potential Chl f-binding site by monitoring changes in the optical properties of variant sr-PSII complexes in which the axial His ligand or the QD residues that are predicted to H-bond to the formyl group are mutated; (4) test the possible involvement of reactive oxygen species in Chl f synthesis (5) establish an in vitro system for Chl f synthesis using either the isolated sr-PSII complex or membranes containing sr-PSII to help assess catalytic parameters of the enzyme and (6) test whether heterologous production of Chl f is enhanced by expressing the native 'far-red' PSII subunits of Chroococcidiopsis thermalis which are known to bind Chl f. In parallel we will (7) isolate FLAG-tagged sr-PSII from C.thermalis to assess whether the native system contains additional protein components and then use mutagenesis to confirm the importance of the QD motif in the native system. Overall this work will provide new insights into the synthesis of Chl f which will be important for future work aiming to introduce Chl f into the photosynthetic apparatus of plants as a strategy to enhance photosynthetic efficiency.

Summary

There is an urgent need to develop new strategies to improve crop yield to feed the ever-growing global population. Crop plants grow because they use the energy of sunlight to drive the conversion of atmospheric carbon dioxide into biomass. This process of photosynthesis is relatively inefficient with much less than 1% of the incident solar energy converted into stored chemical energy. One straightforward way to improve photosynthetic efficiency is to capture more of the sunlight in the first place. Plants rely on chlorophyll pigments (as well as some accessory pigments) to absorb light to drive photosynthesis. The chemical nature of the chlorophyll pigments found in plants necessarily means that photosynthesis is restricted to the visible region of the solar spectrum. In recent years, however, several strains of cyanobacteria, which perform plant-like photosynthesis, have been discovered that make modified forms of chlorophyll that absorb light in the far-red region of the spectrum. If these far-red chlorophylls could be made in plants and assembled correctly in the photosynthetic apparatus, the number of photons of light that could be used to drive photosynthesis could be increased by up to 19%, a considerable increase in efficiency. One of the far-red absorbing chlorophylls is chlorophyll f (Chl f). In order to make Chl f in plants, an important first step is to identify and characterise the cyanobacterial enzyme that synthesises Chl f. In a recent breakthrough, Don Bryant and colleagues in the USA showed that Chl f synthesis was dependent on the ChlF protein subunit which, somewhat surprisingly, was found to be related to one of the proteins present in the well-studied photosystem II complex which catalyses the light-driven oxidation of water to oxygen characteristic of plant photosynthesis. In follow-up work, we have discovered that ChlF does not act alone, as was originally thought, but is part of a new type of PSII complex, which we term the super-rogue PSIIcomplex. The super-rogue PSII complex shows clear similarities to regular PSII but has evolved to make Chl f rather than split water into oxygen. Chl f is made from the Chl a pigment through an oxidation reaction involving molecular oxygen; but the chemistry involved in this process is currently unknown. In this application, we propose to study the structure and mechanism of the newly identified super-rogue PSII complex in unprecedented detail. We aim to investigate whether the super-rogue complex is photochemically active and will test the hypothesis that the super-rogue PSII complex activates molecular oxygen into a reactive form that oxidises a Chl a molecule bound to a specific site in the super-rogue PSII complex. The project involves a team of scientists with skills in microbiology, molecular biology, biochemistry and spectroscopy. Our experimental approaches are diverse and involve working on biochemically pure protein complexes as well studying cyanobacterial mutants expressing Chl f. Ultimately our studies will provide important new knowledge on a new type of photosystem II complex that will underpin future work producing Chl f in crop plants.

Impact Summary

Understanding the details of Chl f synthesis might in the long-term lead to the development of microalgae and plants with more efficient photosynthesis, especially in far-red enriched environments, such as the lower regions of the canopy, or in dense cultures of algae. In the agricultural sector, beneficiaries could include: companies involved in modifying or selecting plants to maintain and or improve crop yields; farmers who wish to develop new practices for similar reasons; governments and policy-makers interested in developing novel strategies to achieve food security; and the public who will benefit from food security. In the biotechnology sector, beneficiaries include companies who wish to develop microalgae and other related photosynthetic organisms as solar biorefineries for the sustainable production of green chemicals and high-value products. In the bioenergy sector, beneficiaries include: companies wishing to develop alternatives to fossil fuels; governments and policy makers who are interested in new routes to energy security and for new energy sources for developing countries; the military who are looking for alternative fuels for specific and niche uses; environmentalists who need to focus on rational long-term alternatives to fossil fuels. In the environmental and ecological sector, beneficiaries include those wishing to understand more about the role of chl f-producing cyanobacteria in a changing environment. In the education sector, in museums and in the media, there will be benefits from publicising new advances in photosynthesis, one of the most fundamental biological processes and one that has been taught at secondary school and so readily familiar to the general public. Staff hired for the project will obtain training in cutting edge research in world-leading research centres. Peter Nixon and James Murray are members of the Photosynthesis Research Lab at Imperial College which includes world leading experts in Photosynthesis including Bill Rutherford FRS and Jasper van Thor, with expertise ranging from femtosecond spectroscopy to cyanobacterial physiology. The PDRA and technical staff will be in an excellent position to progress their careers. They will have the benefit of the excellent intellectual environment of a leading university with a tradition of close ties with engineers and industry.

Committee	Research Committee B (Plants, microbes, food & sustainability)
Research Topics	Structural Biology
Research Priority	X – Research Priority information not available
Research Initiative	X - not in an Initiative
Funding Scheme	X – not Funded via a specific Funding Scheme

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