Supplemental material for: A Glutathione Redox Effect on Photosynthetic Membrane Expression in Rhodospirillum rubrum
Anke Carius, Marius Henkel, and Hartmut Grammel
Application of external GSH to cultivations with Rhodobacter sphaeroides and Rhodobacter capsulatus.Cultivations with external GSH administration were conducted withRhodobacter sphaeroides (ATCC 17023) and Rhodobacter capsulatus (ATCC 17015) both growing in succinate (M2S) medium in shake-flasks.The onset of formation of photosynthetic complexes after the initial aerobic growth phase after 15 h indicates semi-aerobic growth conditions (Supplemental Fig. 1). GSH (1mM) was added after 25 h of cultivation. Photosynthetic membranes were monitored as absorption at 851 nm (R. sphaeroides) and 858 nm (R. capsulatus), respectively using a PowerWave microplate spectrophotometer (Biotek, Winooski, VT). For comparing optical densities (A660 nm values) obtained in 1-cm-path-length cuvettes, the values have to be multiplied with a correction factor of 4.9.
SUPPLEMENTAL FIGURE 1. Growth and formation of photosynthetic complexes in R. sphaeroides and R. capsulatus after stimulation with 1 mM GSH. Symbols: R. sphaeroides: A660 (control), ; A851/A660 (control), ●; A660 (+ 1mM GSH), ▲; A851/A660 (+ 1mM GSH), ■; R. capsulatus: A660 (control), ; A858/A660 (control), ○;A660 (+ 1mM GSH), ∆; A858/A660 (+ 1mM GSH), □. Error bars represent standard deviation of three cultivations.
Total amounts of intracellular GSHwas determined in cell extracts as described in Material and Methods after harvesting of the cells after 53 h and 64 h for R. sphaeroidesand R. capsulatus, respectively. For R. sphaeroides the total concentration of GSH was 4.5 µmol/g CDW (average value of six measurements with standard deviation 0.06) and for R. capsulatus 4.17 µmol/g CDW (average value of six measurements with standard deviation 0.3). In comparison, in R. rubrum, the identical treatment resulted in a final cellular GSH level of 24 µmol/g CDW (see Fig. 2 (B) of the paper).