S31-003 Molecular origin of red pigments in a peripheral light-harvesting antenna of Photosystem I: Ultrafast absorption spectroscopy of recombinant Lhca4 AN Melkozernov 1,2 , S Lin 1,2 , VHR Schmid 3 , E Lago-Places 3 , H Paulsen 3 , and RE Blankenship 1,2 1 Department of Chemistry and Biochemistry and 2 Center for the Study of Early Events in Photosynthesis, Arizona State University, Tempe, AZ, 85287-1604, USA. alexander.melkozernov@asu.edu 3 Institut für Allgemeine Botanik, Johannes Gutenberg-Universität, 55099 Mainz, Germany. vschmid@mail.uni-mainz.de Keywords: LHCI, antenna, femtosecond spectroscopy, energy transfer, low energy chlorophylls Introduction A remarkable feature of Photosystem I in higher plants and green algae are chlorophylls absorbing and emitting at energies lower than P700. A dominant portion of the red fluorescence comes from a peripheral light-harvesting antenna (LHCI), specifically from its LHCI-730 subpopulation. One of the constituents of the LHCI- 730 heterodimer, the Lhca4 subunit, was found to harbor the low energy (red) pigments emitting at 730 nm (Tjus et al. 1995; Schmid et al. 1997; Knoetzel et al. 1998). Excitation energy transfer processes in the LHCI-730 and molecular organization of the red pigments that lead to the excitation energy localization are poorly understood. 77 K transient absorption difference spectra of Lhca4 upon excitation of Chl b revealed the presence of several ultrafast energy transfer processes with yet-unresolved lifetimes (Melkozernov et al. 2000a). Two major energy transfer processes include a 400-600 fs energy transfer between spectral forms of Chl b and Chl a followed by 3-5 ps energy equilibration between Chl a molecules - including those involved in the red pigment’s transition at 705 nm. A significant red shift of the C-705 spectral form as compared to the major Chl a spectral form at 679 nm might be a result of a strong excitonic interaction of the Chl a molecules localizing excitations in the Lhca4. Ihalainen et al. (2000) suggested the presence of one Chl a dimer per LHCI-730 based on steady state spectroscopy of native LHCI complexes. Recently, Schmid et al. (2001) obtained evidence of Chl b involvement in the long-wavelength transition in Lhca4 based on steady state spectroscopy analysis of reconstituted Lhca4. In this work we use ultrafast transient absorption spectroscopy at 8 K to probe coupling of Chl a and Chl b in the recombinant Lhca4 polypeptides with changed pigment occupancy as compared to the wild type Lhca4. Materials and methods Preparation of reconstituted Lhca4. Lhca4 protein overexpression and reconstitution of polypeptides with mixtures of Chl a, Chl b and xanthophylls were described earlier (Schmid et al. 2001). The E102S mutation of the Lhca4 polypeptide was obtained by the PCR based method described by Chen and Przybyla (1994). Isolation of reconstituted Lhca4 (r-Lhca4) by density gradient ultracentrifugation and pigment analysis was as in Schmid et al. (2001).