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The activity of citramalate synthase (CimA, EC 2.3.1.182) (3±0.5 nmole/min·mg protein) and theronine deaminase (60±5 nmole/min·mg protein) can be detected in the cell-free extracts of the minimal medium containing pyruvate.
No differences in the 13C-isotopomer abundances of the protein-derived amino acids for cultures grown in the minimal medium containing pyruvate were observed under either dark or illuminated conditions (Table S1a and S1b).
Spectral features of the photosynthesis system and light-harvesting complexes for the cultures grown in the minimal medium containing pyruvate or glucose are similar to the cultures grown in the rich medium (Figure 1a and 1b) as well as the results reported previously [13], [16].
Moreover, no CO2-enhanced growth was obtained in the minimal medium containing pyruvate or glucose (Figure 1d and 1e), and 13C-isotopomer distribution of protein-based amino acids suggests that the presence of bicarbonate has a rather small effect on pyruvate and glucose metabolism in R. denitrificans (Table S1c and S1f).
Then, 0.1 ml of the pellet was inoculated onto Lowenstein-Jensen medium containing pyruvate or glycerol and incubated for 10 weeks at 37°C [ 27].
To evaluate whether DMPK could have an impact on cell viability by modulating the mitochondrial redox equilibrium, we placed cells in a medium containing pyruvate and glutamine, but devoid of serum and glucose.
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Throughout the QRT-PCR data analysis, we observed that the transcript level for all of the genes tested is higher in cells grown in the minimal medium containing either pyruvate or glucose than in the rich medium (Figure 3).
Requirement for vitamin B12 in the growth of R. denitrificans may partially explain why yeast extract was necessary to be included in the minimal medium containing either pyruvate or glucose.
The QRT-PCR results indicate that both eda (RD1_2878) and edd (RD1_2879) genes are expressed and the transcript level of these genes is higher in the minimal medium containing either pyruvate or glucose than in the rich medium, similar to the gene expression profiles of other genes responsible for carbon fixation and carbon metabolism examined in this report (Figure 3).
After neutralization, sediments were placed on two slants of Löwenstein-Jensen (LJ) medium containing 0.5% pyruvate and one slant containing glycerine.
HEK293 cell culture medium also contained pyruvate.
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