Fig. 6
Allosteric PYK-based molecular OR logic gate synergistically regulates energy and carbon metabolism in mycobacteria. Histograms of metabolic changes at S4, S10, S22 and R6 against Log in nutrient-starvation model; abundance data were normalised to protein concentration and represent mean ± SD, n = 4; significance is indicated as *p < 0.05, **p < 0.01, ***p < 0.001 by one-way ANOVA with Dunnett post test versus Log. a Schematic illustration of glycolysis pathway and metabolic changes in M. bovis BCG during nutrient starvation. The allosteric activator of MtbPYK, glucose-6-P (G6P), is shown in magenta, while PYK substrates (ADP and PEP) and products (ATP and pyruvate) are coloured in red. b A schematic representation of the molecular OR gate. The metabolite G6P (magenta square) and the low-energy-state signal AMP (purple rectangle) are two molecular inputs into the gate which is composed of inactive MtbPYK and its substrates (red trapezoid). The enzyme MtbPYK is activated (output) by the binding of either molecule input or both molecules cooperatively at certain concentrations. The sensitivity to one input molecule is increased as the concentration of the other input molecule increases. The MtbPYK tetramers in inactive T-state, AMP-activated R-state, G6P-activated R-state and AMP/G6P-activated R-state are all shown in schematic representations. Domains are highlighted in one subunit: A domain in green, B domain in grey, C domain in blue. Secondary structures and residues, that undergoes significant movements from T-state to activated R-state, are shown as cartons and sticks. AMP loops are indicated as red lines. A logic gate table is also shown on top. High = high concentration; Low = low concentration. c A three-dimensional (3D) graph shows the relation of activator concentrations and MtbPYK activity in vitro. ‘%V max’ is expressed as the percentage of maximum velocity in the presence of saturating activators. The activities were measured in vitro using purified MtbPYK in the presence of 4 mM ADP and 0.2 mM PEP. d Thermal shift assay results for five MtbPYK complexes. All data are mean ± SEM for two independent experiments done in duplicate
