EPR study on the photo-reduction of spin probe in the green alga Chlorella ellipsoidea

Abstract

CO2 sensing system is believed to exist on the surface of plasmalemma of the green alga Chlorella ellipsoidea. It is well known that C. ellipsoidea can adjust the activity of carbon concentrating mechanism (CCM) in response to growth CO2 concentrations. We prepared two wild-type cells of C. ellipsoidea, i.e., High CO2 (5%)-grown cells and Low CO2-grown cells that have low and high CCM activities, respectively. CO2 insensitive mutants of C. ellipsoidea, which express the maximum level of CCM even grown in 5% CO2, were also cultured under low and high CO2-conditions. Cells were treated with 2 mM of 4-oxo-TEMPO(2,2,6,6-tetramethyl-4-oxopiperidinonxy) (TEMPONE), a spin probe, and 40 mM of K3Fe(CN)6, an oxidant. EPR signals from intracellular TEMPONE were observed under illumination or the dark in the presence of 150 mM of Na2MnEDTA, a signal quencher for external TEMPONE. The signal intensity of intracellular TEMPONE decreased gradually in a time dependent manner even under the dark, but the rate of reduction was stimulated as light intensity was increased. Two mM of DCMU, a specific inhibitor for the electron transport from QA to QB, halted the reduction of EPR signal. These data indicate that intracellular TEMPONE is reduced by photosynthetic electron flow primarily at PS II. Furthermore, the difference in effects of wavelength of light on the photoreduction of TEMPONE were also investigated.