All MD simulations were performed under isothermic-isobaric conditions at 298 K using the GROMOS package of programs (48). The atomistic GROMOS force field 54A7 (49) was used with the simple point charge (SPC) model as water model. Initial parameters for SAM and SAH were taken from the Automated Topology Builder server (50) and adjusted to match the parameters of related substructures in amino acids. The bond lengths were constrained to the ideal values applying the SHAKE algorithm (51). The temperature was maintained close to its reference value T = 298 K by weak coupling to a temperature bath with a relaxation time of 0.1 ps (52). The pressure was maintained close to its reference value P = 1.013 bar (1 atm) by weak coupling to a pressure bath with a relaxation time of 0.5 ps and using the isothermal compressibility kT = 4.575 × 10−4 (kJ mol−1 nm−3)−1. Newton’s equations of motion were integrated using the leap-frog scheme (53) with a time step of 2 fs. A reaction field force (54) was applied using the relative dielectric permittivity εrf = 61(55). As initial coordinates, the crystal structure of OphAΔC6 with SAM was taken. All crystal waters were removed with the exception of one water molecule buried in each binding pocket. The two magnesium cations were included. The missing loop between residues 268 and 277 in monomer A was modeled using the webserver ModLoop (56). The missing loops between residues 380 and 405 in monomer A and between residues 379 and 404 in monomer B were not modeled due to their length, that is, the substrates in the binding pockets were not connected to the parent proteins. The dimer complex was energy-minimized in vacuum and solvated in a cubic box of 21516 SPC water molecules. The solvent was energy-minimized with the protein position restrained. A thermalization in five steps from 60 to 298 K was performed with the position restraints decreased from 2.5 × 104 kJ mol−1 nm−2 to zero, with the first four steps being 20 ps under constant-volume condition and the last step being 500 ps under isothermic-isobaric condition. A production run of 20 ns was performed. The simulations were analyzed in terms of backbone atom-positional rmsd and secondary structure motifs using the GROMOS++ analysis programs (fig. S8) (57). As QM calculations require a defined location of the proton in the product state for convergence, a snapshot where the water molecules formed a hydrogen-bonding network up to the deprotonated carboxy group of SAM was taken from the MD simulation as a starting point. If necessary, the orientation of key residues in the binding pocket was modified using PyMol (Schrödinger LLC) to obtain a hydrogen-bonding network similar to that observed in the crystal structure of OphAΔC6. The final geometry is shown in Fig. 5B.

All electronic structure calculations were performed with the quantum chemistry software package Gaussian09 Rev. D1 (Gaussian Inc.), using Kohn-Sham density functional theory (58). B3LYP (59, 60) and TPSSH (61) were used as exchange correlation functional. The orbitals were expanded in the 6-31G** basis set (62, 63) in conjunction with density fitting for the two-electron integrals (64, 65). The size of the integration grid was chosen as UltraFine. To account for solvation effects, a reaction field within the integral equation formalism model (66) was applied. As solvent, diethylether with a dielectric constant of ε = 4.24 was chosen, which is a common approximation for the surrounding enzyme environment (67). For the minimization of the electronic energy with respect to the nuclear coordinates and the location of the transition state, atoms far away from the reaction center were kept frozen. More specifically, the methyl group of SAM was chosen as center, and all atoms with a distance >7 Å to it were constrained (fig. S8). Once a saddle point was located, a unique and coordinate-independent reaction coordinate, the so-called intrinsic reaction coordinate (IRC), was followed. By definition, this corresponds to an imaginary minimum energy trajectory in mass-weighted Cartesian coordinates with the initial direction indicated by the normal mode of the imaginary frequency of the transition state (68). In essence, this trajectory corresponds to the steepest decent path or minimum energy path (69, 70). The IRCs of step 1 and 2 are shown in fig. S8.

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