Circularly Polarised Luminescence Spectroscopy of Photosystem II

Abstract

An improved understanding of the fluorescent excited states of chlorophyll in Photosystem II (PSII) is needed for several aspects of photosynthesis research. This thesis describes an investigation into these excited states, carried out using circularly polarised luminescence (CPL) spectroscopy. The CPL signal of photosynthetic proteins arises due to exciton interactions between chlorophyll molecules within the enzyme, in a similar manner to the better-known circular dichroism (CD) technique. Measurements were carried out at low temperature on purified preparations of the PSII core complex and two of its substituents, the proximal antenna proteins CP43 and CP47. Development of the low-temperature CPL measurement technique was a major part of the work. This was necessary to demonstrate that polarisation artifacts were small, and hence to ensure that circular polarisation was measureable with the required precision (within 1 part in 10^4). The utility of temperature-dependent CPL measurements in the range of 2– 120 K was demonstrated on the already well-characterised CP43 protein. The lowest-energy excited states of this protein have negative CPL, which matches their exciton CD as expected. This result does not require any change to prior assignments of the low-energy states of CP43. There are significant open questions regarding the lowest-energy excited state of the CP47 protein. Careful measurements on this protein reveal a positive CPL signal for this state (wavelength ≥690 nm), which also matches its CD spectrum. Using structure-based exciton calculations, it is proposed that the lowest state of CP47 is most likely located on Chlorophyll 612. This is a significant departure from the previous assignment of Chlorophyll 627. Measurements on intact PSII core complexes confirm that the same low-energy state is present in these complexes as in CP47, resulting in a new assignment for the well-known F695 fluorescence band. The new assignment is found to be consistent with many aspects of the published PSII literature. Show more