Table 1.

The effect of metabolic regulators and monovalent ions on PykA activity

Kinetic parameters were calculated using GraphPad Prism from best-fit nonlinear regression analysis of the data. The allosteric sigmoid function was used for the PEP titration and the Michaelis-Menten function was used for ADP titration.

	No additive	0.2 mm F6P	0.15 mm R5P	1 mm G3P	1 mm G6P	1 mm KDPG	0.5 mm X5P	1 mm RL5P	100 mm KCl	100 mm NH4Cl	100 mm NaCl
PEP titration
s0.5 (mm)	0.670 ± 0.058	0.159 ± 0.021	0.195 ± 0.017	0.297 ± 0.017	0.290 ± 0.026	0.432 ± 0.036	0.209 ± 0.011	0.269 ± 0.012	1.187 ± 0.086	1.402 ± 0.084	1.492 ± 0.116
Hill coefficient (h)	2.14 ± 0.34	1.45 ± 0.31	1.65 ± 0.25	1.89 ± 0.21	1.78 ± 0.32	1.52 ± 0.18	1.87 ± 0.22	1.61 ± 0.12	2.36 ± 0.28	2.31 ± 0.22	2.42 ± 0.32
Vmax (mm/min)	0.099 ± 0.004	0.096 ± 0.004	0.092 ± 0.003	0.092 ± 0.002	0.100 ± 0.003	0.100 ± 0.004	0.110 ± 0.002	0.110 ± 0.002	0.087 ± 0.002	0.130 ± 0.003	0.120 ± 0.004
kcat(s−1)	431.9	418.8	401.4	401.4	436.3	436.3	479.9	479.9	379.6	567.2	523.6
kcat/s0.5(s−1 mm−1)	644.1	2626	2055	1348	1504	1008	2286	1778	319.8	404.6	350.9
ADP titration
Km (mm)	0.070 ± 0.015	NAa	NA	NA	NA	NA	NA	NA	0.147 ± 0.020	0.155 ± 0.026	0.146 ± 0.033
Vmax (mm/min)	0.080 ± 0.005	NA	NA	NA	NA	NA	NA	NA	0.089 ± 0.004	0.067 ± 0.003	0.078 ± 0.005
kcat (s−1)	349.0	NA	NA	NA	NA	NA	NA	NA	388.3	292.3	340.3
kcat/Km (s−1 mm−1)	4952	NA	NA	NA	NA	NA	NA	NA	2633	1876	2321

^a NA, not applicable (the regulator had no significant effect on PykA kinetics compared with the control (no additive present)). The values of k_cat were calculated using [E_t] = [PK monomer].