Femtosecond Visible/Visible and Visible/Mid-IR Pump-Probe Study of the Photosystem II Core Antenna Complex CP47 Marie Louise Groot,* ,² Jacques Breton, ‡ Luuk J. G. W. van Wilderen, ² Jan P. Dekker, ² and Rienk van Grondelle ² Faculty of Sciences, Vrije UniVersiteit, 1081 HV Amsterdam, The Netherlands, and SerVice de Bioe´ nerge´ tique, Baˆ t. 532, CEA-Saclay, 91191 Gif-sur-YVette, France ReceiVed: December 22, 2003; In Final Form: March 16, 2004 CP47 is one of the two core antenna proteins of Photosystem II involved in the transfer of solar energy toward the photochemically active reaction center, the D1D2cytb559 complex. We have performed vis/vis and vis/mid-IR pump-probe experiments at room temperature as a first step in linking the energy-transfer dynamics to the arrangement of the individual chlorophylls in the CP47 complex. The chlorophylls in CP47 have very similar absorption maxima (within 20 nm of each other); therefore, few spectral changes due to energy transfer can be observed at room temperature. We used the annihilation of excitation energy as a tool to enhance the spectral changes associated with energy transfer. Energy transfer was found to occur on time scales of ∼100 fs, 1-4 ps, and 12-28 ps, and our results are consistent with the presence of two red (683- nm) pools, plus an additional red-shifted one (690 nm). From the time-resolved mid-IR spectra, it follows that these red states show a keto CdO stretching frequency at 1686 cm -1 and therefore are either in a polar environment or have a fairly weak hydrogen bond. For the more blue-absorbing states, we observe varying keto band positions between 1696 and 1664 cm -1 , and thus their hydrogen bonds strength varies between “none present” and “strong”. A change in the frequency of the coordination marker mode was observed when the 690-nm state was populated, probably caused by a more planar conformation of the macrocyle of the chlorophyll responsible for the 690-nm state. 1. Introduction The primary steps in photosynthesis of energy and electron transfer occur in green plants in two large protein complexes called Photosystem I and Photosystem II. The Photosystem II core complex consists of several individual pigment-protein complexes; these are the core antenna’s CP43 and CP47 and the D1D2cytb559 reaction center. Recently, two crystal struc- tures of PSII from cyanobacteria were reported, one at 3.8-Å 1 and one at 3.7-Å 2 resolution. In the latter structure, 17 Chl a molecules were found in CP47, which is one more than could be observed in the earlier structure. CP47 also binds two or three -carotenes that were not observed in the structure. 1,2 The chlorophylls are roughly distributed in two layers near the stromal and lumenal sides of the membrane. Most of the pigments are oriented with their plane perpendicular to the membrane plane. The resolution with which these crystal structures are resolved is not high enough to recognize some of the details of the cofactors, such as the orientation of the ring plane or their binding with the protein. 23 The CP47 and CP43 complexes serve to absorb solar photons and transfer the excited-state energy to the D1D2 reaction center. The excitation-energy trapping time in PSII cores with open RCs is about 50-100 ps; 3-5 in cores of plant PSI, with close to 200 Chls, trapping times of 50 and 120 ps have also been measured. 6 Energy transfer between groups of pigments within the isolated CP47 complex occurs on the 0.2-0.4 ps, 2-3 ps, and ∼20 ps time scales at 77 K. 7 Pools of pigments absorbing at 660, 670, 676, and 683 nm can be recognized from the (low- temperature) absorption spectrum. 7-9 An exciton calculation of the chlorophylls based on the published structure yielded results that were in agreement with the absorption spectrum and were in line with the observed energy-transfer rates. 7 From these calculations, it followed that the pigments are mainly involved in pairwise interactions. Both on the lumenal and on the stromal side a pair of Chls (35-48 and 39-42 in the Zouni nomen- clature 1 ) were suggested to give rise to the lowest exciton band at 683 nm. There is experimental evidence that a 690-nm state also exists in CP47, from linear dichroism, 8 hole-burning studies, 10,11 and fluorescence line-narrowing studies. 12 From a simultaneous fit of the linear dichroism and absorption spectrum, it was concluded that this state has the oscillator strength of one Chl and an ∼20% larger width than the other Chl pools (217 cm -1 at 77 K). 12 A weak band at 1633 cm -1 observed in the fluorescence line-narrowed spectrum was assigned to the Chl keto band and interpreted to be indicative of a very strong hydrogen bond with the protein, thus possibly causing the red- shifted absorption of this Chl. 12 In the present study, we report the results of femtosecond visible pump/mid-IR probe experiments on CP47 detected in the 1600-1800-cm -1 region. This region mainly probes the absorption changes in the CdO stretches of the chromophore (i.e., the 9-keto and 10a-ester modes) and the protein, and a mode sensitive to the macrocycle of the chromophore. The keto and ester modes are sensitive to the polarity of the environment and the presence and strength of hydrogen bonds that the Chl may engage at these positions with the protein. Energy transfer between pigments in different environments or with different * Corresponding author. E-mail: ML.Groot@few.vu.nl. ² Vrije Universiteit. ‡ Service de Bioe´nerge´tique. 8001 J. Phys. Chem. B 2004, 108, 8001-8006 10.1021/jp037966s CCC: $27.50 © 2004 American Chemical Society Published on Web 05/14/2004