Fig. 2: Biochemical characterization of OGDHc.

A Western blots displaying the detection of all OGDHc components in the native cell extract fraction at the expected molecular weight (MW). A low-MW fraction was used as negative control, whereas the overexpressed protein that was used to create the antibodies was employed as a positive control. Due to the large size of the E1o protein, a fragment was employed for this reason (see Methods). Corresponding bands have been annotated with an orange arrow. B Enzymatic characterization of the native OGDHc. α-Ketoglutarate, NAD⁺ and CoA were used at the concentrations shown in each plot accordingly, the velocity was normalized to 0-1 and a line is connecting each point, with different symbols annotating the data points for the three independent biological replicates, as annotated in the figure panel. C Graph displaying the change in reaction velocity (after normalization to 0-1) in relation to temperature for C. thermophilum as compared to a yeast equivalent sample, with different symbols annotating the data points for the three independent biological replicates, as annotated in the figure panel. The K_M values shown in plots B and C were obtained by the Burk-Lineweaver plots shown in Supplementary Fig. 2B and the gray background for each graph and the black bars at the bottom graph represent the standard deviation derived from N = 3 independent biological replicates and 2 technical duplicates for each replicate. All values shown here are listed in Supplementary Data 1.