Spectral, Photophysical, and Stability Properties of Isolated Photosystem II Reaction Center

Photosystem II reaction center (RC) preparations isolated from spinach (Spinacea olerácea) by the Nanba-Satoh procedure (O Nanba,K Satoh 1987 Proc Natl Acad Sci USA 84: 109-112) are quite labile, even at 4°C in the dark. Simple spectroscopic criteria were developed to characterize the native state of the material. Degradation of the RC results in (a) blue-shifting of the red-most absorption maximum, (b) a shift of the 77K fluorescence maximum from ~682 nm to ~670 nm, and (c) a shift of fluorescence lifetime components from 1.3-4 nanoseconds and >25 nanoseconds to~6-7 nanoseconds. Fluorescence properties at 77K seem to be a more sensitive spectral indicator of the integrity of the material. The >25 nanosecond lifetime component is assigned to P680⁺ Pheophytin^-recombination luminescence, which suggests a correlation between the observed spectral shifts and the photochemical competence of the preparation. Substitution of lauryl maltoside for Triton X-100 immediately after RC isolation stabilizes the RCs and suggests that Triton may be responsible for the instability.