Several proteins within the glycolysis and pyruvate metabolism pathways were up-regulated (e.g Gap1, Pgi), whereas all glycolytic genes except eno were up-regulated in the microarray experiment.
To measure the effect of metabolic mutations on carbon flux polarity, competent cells of the GM1514 strain that contains lacZ downstream from the gapB promoter at the amyE locus were transformed with the total DNA extracted from strains carrying metabolic mutations (168 Pgi::spc, 501 77, GM1501, GTD040 and DGRM18, 25).
An increase of the glucose uptake rate by overexpression of the glucose facilitator gene glf, and the glucokinase gene glk from Zymomonas mobilis resulted in a higher specific biotransformation rate in the strains SS02 (pgi deletion) and SS03 (pfkA deletion).
A fourth gene, YPO3718 (pgi), is predicted to be essential on gluconate only because its deletion disrupts gluconeogenesis and consequently the biosynthesis of glycogen, an indispensable component of the YP CO92 biomass objective function.
WS-4 is WS wspR::mini-Tn5; WS-12 is WS pgi::mini-Tn5, and WS-13 is WS wssB::mini-Tn5 [2].
Likewise, plasmid p∆ ppc was constructed using K972 + K973 and K974 + K975, plasmid p∆ aceE was constructed using K960 + K961 and K962 + K963, and plasmid p∆ pgi with primers K950 + K951 and K952 + K953 and plasmid p∆ ilvE was constructed using K21 + K22 and K23 + K24.
It is widely perceived that plants do not flower even under photo-inductive conditions until they accumulate enough sugar reserves for the induction of flowering [ 6- 8], which is consistent with the observations that low-starch-containing mutants, such as pgm1 and pgi, exhibit retarded growth and delayed flowering [ 9, 10].
The specific glucose uptake rate was very high for the reference strain (11.3 mmol h−1 gcdw−1) but in the range of previously reported values (Toya et al. 2010) and significantly lower for the Δ pfkA mutant (4.0 mmol h−1 gcdw−1) and the Δ pgi mutant (2.7 mmol h−1 gcdw−1).
Obviously, at this early time point after the pulse the G6P converting enzymes, Pgi and G6pdh, are the most limiting steps for the further conversion of the glucose after uptake.
ET-1, PGI-2, and NO concentrations in the supernatant were measured using ELISA kits according to the manufacturer's instructions, respectively.
The concentrations of ET-1, PGI-2, and NO in the supernatant were measured by enzyme-linked immunosorbent assay (ELISA) kits (Jiancheng, Nanjing, China), respectively.
