Table 2.

Simulation and experimental analysis of Gibbs energy activation of TS formation of WTIgP and its variants in reaction with POX from substrate complex to TS

	ΔG‡,* kcal/mol	ΔG‡Cζ,† kcal/mol	ΔG‡Cβ,§ kcal/mol
l-Ser35Arg	21.2 ± 0.3	21.5	21.3
l-Ser35Lys	21.1 ± 0.4	25.7	23.8
l-Leu47Arg	20.9 ± 0.8	30.0	25.3
l-Leu47Lys	21.4 ± 0.7	24.6	21.3
WTIgP	ND¶	28.5	29.3

^*ΔG^‡, Gibbs energy of activation calculated using Eyring–Polanyi equation.

^†ΔG^‡_Cζ, Gibbs free energy for TS formation calculated by combined QM/MM and funnel metadynamics using the following constraints: dTyrO–P = 1.5 to 4 Å; angle Tyr_Cζ–P–O_4-NP = 145 to 180°.

^§ΔG^‡_Cβ, Gibbs free energy for TS formation calculated by combined QM/MM and funnel metadynamics using the following constrains: dTyrC_β–P = 6 to 9 Å; angle Tyr_Cβ–P–O_4-NP = 120 to 180°.

^¶ND, not determined because of low rate of reaction.