The remarkable near-unity quantum efficiency of photosynthetic charge separation has motivated decades of research to uncover the underlying design principles. Much of our current understanding of photosynthetic charge separation is rooted in studies of the bacterial reaction center (BRC). We present two-dimensional electronic spectroscopy of the BRC as it undergoes charge separation, resolving the energy-transfer and charge-separation processes with time and excitation frequency resolution. These measurements reveal the excitonic structure of the BRC, including a previously hidden exciton state. We present a multiexcitation 2D global analysis method that supports two-step sequential charge separation in the BRC without evidence for secondary charge separation pathways. We extract the spectral signatures of the charge-separation intermediates.
In the initial steps of photosynthesis, reaction centers convert solar energy to stable charge-separated states with near-unity quantum efficiency. The reaction center from purple bacteria remains an important model system for probing the structure–function relationship and understanding mechanisms of photosynthetic charge separation. Here we perform 2D electronic spectroscopy (2DES) on bacterial reaction centers (BRCs) from two mutants of the purple bacterium Rhodobacter capsulatus, spanning the Q y absorption bands of the BRC. We analyze the 2DES data using a multiexcitation global-fitting approach that employs a common set of basis spectra for all excitation frequencies, incorporating inputs from the linear absorption spectrum and the BRC structure. We extract the exciton energies, resolving the previously hidden upper exciton state of the special pair. We show that the time-dependent 2DES data are well-represented by a two-step sequential reaction scheme in which charge separation proceeds from the excited state of the special pair (P*) to P +H A − via the intermediate P +B A −. When inhomogeneous broadening and Stark shifts of the B* band are taken into account we can adequately describe the 2DES data without the need to introduce a second charge-separation pathway originating from the excited state of the monomeric bacteriochlorophyll B A*.