Figure 2.

Analysis of pigments in Synechocystis mutant strains. A, Quantification of the relative contents of PP_IX, MgP, magnesium protoporphyrin IX methylester (MgPME), PChlide, and Chlide in Synechocystis cells growing at normal light (40 μmol photons m⁻² s⁻¹) and harvested at OD₇₅₀ = 0.35 to 0.4. Chl precursors were extracted with methanol/0.2% NH₄OH and quantified by a combination of HPLC and spectrofluorometry. Values shown represent means ± sd from three independent measurements; asterisks indicate significant differences tested using a paired t test (P = 0.05). B, ALA-synthesizing capacity as determined in a 100-mL cell suspension with OD₇₅₀ = 0.4 treated for 4 h with levulinic acid to inhibit ALA condensation into porphobilinogen. Representative absorption spectra of developed assays as obtained for each strain together with 5 μm standard of ALA are also presented. C, Quantification of noncovalently bound heme B (protoheme) in cells growing at normal light. Heme was extracted by 90% acetone/2% HCl, separated, and quantified using HPLC. Error bars and the t test of statistical significance are as in A. D, Quantification of the relative contents of Chl precursors in Synechocystis cells growing for approximately 40 h at high light (200 μmol photons m⁻² s⁻¹) and harvested at OD₇₅₀ = 0.35 to 0.4. Obtained values from three measurements were compared with values for the control WT_zeo strain growing at normal light. The dashed line indicates 100% in comparison with values at normal light (A). Asterisks indicate statistically significant differences in precursor levels between WT_zeo and ΔH347 at high light as tested using a paired t test (P = 0.05).