Biophysical Chemistry 106 (2003) 221–231 0301-4622/03/$ - see front matter  2003 Elsevier B.V. All rights reserved. doi:10.1016/S0301-4622(03)00215-1 Excited state charge-transfer dynamics study of poplar plastocyanin by ultrafast pump-probe spectroscopy and molecular dynamics simulation Tiziana Cimei , Anna Rita Bizzarri , Giulio Cerullo , Sandro De Silvestri , a a b b Salvatore Cannistraro * a, Biophysics & Nanoscience Group, INFM, Dipartimento di Scienze Ambientali, Universita della Tuscia, I-01100 Viterbo, Italy a ` National Laboratory for Ultrafast and Ultraintense Optical Science-INFM, Dipartimento di Fisica, Politecnico di Milano, b Piazza Leonardo da Vinci 32, I-20133 Milano, Italy Received 3 July 2003; received in revised form 18 July 2003; accepted 22 July 2003 Abstract We have applied ultrafast pump-probe spectroscopy to investigate the excited state dynamics of the blue copper protein poplar plastocyanin, by exciting in the blue side of its 600-nm absorption band. The decay of the charge- transfer excited state occurs exponentially with a time constant of approximately 280 fs and is modulated by well visible oscillations. The Fourier transform of the oscillatory component, besides providing most of the vibrational modes found by conventional resonance Raman, presents additional bands in the low frequency region modes, which are reminiscent of collective motions of biological relevance. Notably, a high frequency mode at approximately 508 cm , whose dynamics are consistent with that of the excited state and already observed for other blue copper y1 proteins, is shown to be present also in poplar plastocyanin. This vibrational mode is reproduced by a molecular dynamics simulation involving the excited state of the copper site.  2003 Elsevier B.V. All rights reserved. Keywords: Poplar plastocyanin; Ultrafast pump-probe spectroscopy; Charge-transfer excited state; High frequency mode; Duschin- sky rotation; Molecular dynamics simulation 1. Introduction The application of femtosecond pump-probe spectroscopy to study the ligand-to-metal charge- transfer (LMCT) dynamics of several blue copper proteins w1–4x, has very recently gained large *Corresponding author. Tel.: q39-076-135-7136; fax: q39- 076-135-7136. E-mail address: cannistr@unitus.it (S. Cannistraro). popularity, particularly because it gives informa- tion on the excited state nuclear dynamics w2,5– 8x and in addition circumvents the experimental difficulties of Resonance Raman (RR) spectro- scopy in discriminating the low frequency modes w5x. Moreover, the dynamics following the optical preparation of the LMCT band may provide useful information on the processes associated with the thermal electron transfer (ET) reaction in blue proteins, even if the charge-transfer that occurs is