Wavelength and Intensity Dependent -z VED 1996 T I

Wavelength and Intensity Dependent

Reaction Centers Using an Optical Parametric Amplifier ’ -z VED

2 ? 1996 Scott R. Greenfield, Chemistry Division, Argonne National Laboratory, Argonne, I l l inoio s T I

60439-4831, (708) 252-3541

Michael Seibert, Basic Sciences Division, National Renewable Energy Laboratory, Golden, Colorado 80401, (303) 384-6279

Govindjee, Department of Plant Biology, University of Illinois, Urbana, Illinois 61801- 3707, (217) 333-1794

Michael R. Wasielewski, Chemistry Division, Argonne National Laboratory, Axgome, Illinois 60439-483 1, (708) 252-3538, and Department of Chemistry, Northwestern University, Evanston, Illinois 60208-3 113, (708) 467-1423

Abstract

Isolated Photosystem I1 reaction centers were excited at five wavelengths to study the

effects of excitation wavelength and intensity on energy transfer and charge separation.

DISCLAIMJ3R

This report was prepared as an account of work sponsored by an agency of the United States Government. Neither the United States Government nor any agency thereof. nor any of their employees, makes any warranty, express or implied, or assumes any legal liability or responsi- bility for the accuracy, completeness, or usefulness of any information, apparatus, product, or process disclosed, or represents that its use would not infringe privately owned rights. Refer- ence herein to any specific commercial product, process. or service by trade name, trademark, manufacturer, or otherwise does not necessarily constitute or imply its endorsement, recom- mendation, or favoring by the United States Government or any agency thereof. The views and opinions of authors expressed herein do not necessarily state or reflect those of the United States Government or any agency thereof.

The submitted manuscript has k e n authored by a contractor of the U.S. Government under contract No. W-31-104ENG-38. Accordingly, the U. S. Government retains a nonexclusive, royalty-free license to publish or reproduce the published form of this contribution, or allow others to do so, for U. S. Government purposes.

Wavelength and Intensity Dependent Studies of Isolated Photosystem 11 Reaction Centers Using an Optical Parametric Amplifier

Scott R. Greenfield, a Michael Seibert, Govindjee, and Michael R. WasielewskP’d ‘Chemistry Div., Argonne National Lab., Argonne, Illinois 60439-483 1; bas ic Sciences Div., National Renewable Energy Lab., Golden, Colorado 80401; ‘Dept. of Plant Biology, University of Illinois, Urbana, Illinois 61 801-3707; dDept. of Chemistry, Northwestern University, Evanston, Illinois 60208-3 113

The photosystem I1 reaction center (PSII RC) is the site for primary charge separation in oxygenic photosynthesis. The lowest energy singlet states of the six chlorophyll a (Chl) and two pheophytin a (Pheo) pigments found in the PSII RC form the composite Q, band at -675 nm. The long wavelength side of the band (the “red” pigment pool) is dominated by the primary donor, P680 (a dimer or multimer of Chl), while the short wavelength side of the band (the “blue” pigment pool) is composed of the accessory Chl. Energy transfer, occurring on both subpicosecond [ 13 and tens of picoseconds [2] time scales, links the various pigments to P680. The charge separated state, P680+-Pheo-, lacks distinctive spectral features, making it difficult to distinguish between charge separation and energy transfer. Arguably the best indicator of charge separation is the bleach of the Pheo Q, band at 544 nm.

We have undertaken a new transient absorption study that utilizes an optical parametric amplifier (OPA) as a pump source. The two-stage OPA is pumped by the second harmonic of a regeneratively amplified Ti:sapphire laser system. The first stage of the device is a type I1 BBO crystal seeded by a white light continuum, and provides near-transform-limited spectral bandwidths over the entire tuning range (475-825 nm) [3]. These narrow bandwidths (-6 nm) allow us to investigate the pump-wavelength dependence of the kinetics across the composite Qy band with G O O fs time resolution. The polarization of the probe beam was at the magic angle relative to that of the pump beam. Pump and probe spot sizes were -250 pm (diameter). The sample, which was isolated from spinach, was kept at a temperature of 5 “C.

Transient absorption kinetics probing at 544 nm were measured for pump wavelengths of 665 and 683 nm for pump energies ranging from 40 nJ to 1.0 pJ. The data are fit to a triple exponential decay (two exponentials did not produce satisfactory residuals) and a component that does not decay on the sub-nanosecond timescale of the experiment. Typical TS of the triple exponential decay are 1-3 ps, 10-25 ps, and 60-120 ps, with the lower end of the ranges corresponding to 683-nm excitation, and the upper end to 665-nm excitation. The intermediate component has the greatest amplitude (47%5% and 60*4% for 683 and 665-nm excitation, respectively). Neither the 7s nor the relative amplitudes of the three components changed over the investigated pump intensity range. However, the nondecaying component, which reflects the filly charge separated state of the RC, saturated at the highest pump energy densities.

Figure 1A shows transient spectra at a delay of 500 ps. Within the signal to noise of the data, the spectra with 665 and 683 nm excitation wavelengths are identical over the entire region. Thus, after energy transfer and charge separation are complete, the RCs are spectrally indistinguishable. This shows that energy transfer from the “blue” pool to P680 occurs with near unity efficiency, as the quantum yield for charge separation is not effected by which pigment pool is excited.

The transient absorption spectra at 1.0 ps for 665 and 683-nm pump wavelengths are shown in Fig. lB, along with the 665-nm minus 683-nm difference spectrum (on the same scale).

500 520 540 560 580 600 500 520 540 560 580 600 Wavelength (nm) Wavelength (nm)

Figure 1: Transient absorption spectra of isolated the PSII RC with 665 and 683-nm excitation wavelengths. A) At 500 ps. B) At 1.0 ps. Also shown is the 665 minus 683-nm difference spectrum, along with the excited state spectrum of Chl in THF.

40 ln n m 5 30

e e

- u)

9 20

ft 10

01 ’ ’ ’ I 0.0 0.5 1.0 1.5 2.0 2.5 3.0 3.5

Absorbed photonslRC

Figure 2: Zero-crossing times in PSII RCs with a probe wavelength of 544 nm.

This is the first observation of a clear difference in the transient spectra in this wavelength region at early time comparing selective excitation of the two pigment pools. The magnitude of the difference spectrum calls into question the extent of subpicosecond energy transfer. Also shown in Fig. 1B is the transient absorption spectrum of Chl in THF at early time. The features of the PSII difference spectrum are very similar to those of in vitro Chl, indicating that the state from which energy transfer takes place in the PSII RC is an excited state of Chl. The decay of the difference spectrum with time is an indication of energy transfer, and analysis of kinetic traces at 544 nm indicates that almost 90% of the amplitude of the -

difference in the decays for the two pump wavelengths disappears with a 29-ps time constant. Figure 2 shows the time at which the transient absorption decay at 544 nm crosses AA=O

for several pump wavelengths as a function of absorbed photons per RC. The “zero-crossing times” (ZCT) for the two pump wavelengths that selectively excite the “blue” pigment pool clearly fall on the same line. A similar behavior is seen with “red” pool excitation, with the ZCT roughly a factor of two shorter than for “blue” pool excitation. The ZCT is a useful monitor of a number of factors, including the pump energy density, the identity of the pigment pool that has been excited, and sample activity.

This work was supported by the Divisions of Energy Biosciences and Chemicai Sciences, Office of Basic Energy Sciences, U.S. DOE under contracts DE-AC36-83CH10093 (M.S.) and W-3 1-109-Eng-38 (M.R.W.).

References [l]

[2]

[3]

J.R. Durrant, G. Hastings, D.M. Joseph, J. Barber, G. Porter and D.R. Klug, Proc. Natl. Acad. Sci. USA 89 (1992) 11632. T. Rech, J.R. Durrant, D.M. Joseph, J. Barber, G. Porter and D.R. Klug, Biochemishy 33 (1994) 14768. S.R. Greenfield and M.R. Wasielewski, Opt. Lett. 20 (1995) 1394.