Ultrafast resonance energy transfer from a site-specifically attached fluorescent chromophore reveals the folding of the N-terminal domain of CP29

• Published in: Chemical physics 2009, Vol.357, p.1-3
• Main Authors: Oort, B.F., van, Murali, S, Wientjes, E, Koehorst, R.B.M, Spruijt, R.B, Hoek, A., van, Croce, R, Amerongen, H., van
• Format: Article
• Subjects: absorption properties; chlorophyll a/b complex; escherichia-coli; excited-states; green plants; light-harvesting complexes; photosystem-ii subunit; plant antenna protein; pump-probe; refractive-index
• ISSN: 0301-0104

The photosynthetic minor antenna complex CP29 of higher plants was singly mutated, overexpressed in Escherichia coli, selectively labeled with the fluorescent dye TAMRA at three positions in the N-terminal domain, and reconstituted with its natural pigments. Picosecond fluorescence experiments revealed rapid excitation energy transfer (20 ps) from TAMRA covalently attached to a cysteine at either position 4 or 97 (near the beginning and end of the N-terminal domain) to the chlorophylls in the hydrophobic part of the protein. This indicates that the N-terminus is folded back on the hydrophobic core. In 20% of the complexes, efficient transfer was lacking, indicating that the N-terminus can adopt different conformations. Time-resolved polarized fluorescence measurements demonstrate that the non-transferring conformations only allow restricted rotational motion of the dye molecule. When TAMRA was attached to a cysteine at position 40, the overall transfer efficiency was far lower, reflecting a larger distance to the hydrophobic region