64
T. Yumura et al.
rebound trajectories, Friesner et al. estimated 69–84% retention of stereochemistry
of for the ethane hydroxylation on MMO [80] being close to the corresponding
experimental value (72%) [40].
Furthermore, Friesner group used QM/MM calculations to incorporate the entire
protein environment of sMMO to improve potential energy surfaces of the hydroxylation of several substrates (CH 4 , C 2 H 6 , CH 3 OH, CH 3 CN, and CH 3 NO 2 ) [87].
The QM/MM calculations investigated the energetics of the initial binding of the
substrate into the enzyme, as well as that of the sequent C–H bond dissociation. In
these calculations, van der Waals and electrostatic interactions between the protein
and a substrate can be directly accounted to accurately calculate the activation barriers
for both steps. The QM/MM studies found that the energy barrier for the substrate
diffusion is not negligible compared with that in the C–H bond activation. As a result,
Friesner et al. suggested that the energetic information in the catalytic reactions by
MMOH Q , including the substrate binding and the methane activation, is necessary
to understand the experimental findings on kinetics in the hydroxylation reactions.
6 Mechanism for Methane Hydroxylation on pMMO
After obtaining the X-ray crystal structure of the particulate form of MMO (pMMO)
in 2.8 Å resolution [10], Shiota and Yoshizawa performed DFT and QM/MM calculations to elucidate its catalytic function for the methane hydroxylation [114–117].
Taking the crystal structure of pMMO into account, mononuclear and dinuclear
copper species, located within the soluble regions of the pmoB subunit, were considered as the active site for the methane to methanol conversion [114]. A reaction
of the mononuclear site with O 2 results in the formation of a Cu(III)–O species in
an endothermic manner, and that of dinuclear site leads to a (μ-O) 2 Cu(II)Cu(III)
species in an exothermic manner. The optimized structures for the two active sites
surrounded by the protein environment were obtained in the QM/MM methods, as
shown in Fig. 13. Furthermore, Shiota and Yoshizawa assumed that a (μ-O)(μOH)Cu(II)Cu(III) species is formed as an active site of pMMO by the H-atom
transfer from Tyr374 residue outside the first coordination sphere to assist the activation of the O–O bond [116]. The formation of the (μ-O)(μ-OH)Cu(II)Cu(III)
active site was proposed by an analogy to the oxidation of tyrosine to dopaquinone
by tyrosinase [122]. On the other hand, Chen et al. proposed a (μ 3 -O)trinuclear
Cu(II)Cu(II)Cu(III) active site for the methane hydroxylation, which proceeds via a
concerted oxo-insertion manner [123, 124]. However, such a trinuclear copper site
was not observed in the X-ray crystal structure of pMMO [10].
Here, let us focus on mechanisms for the methane hydroxylation by the Cu(III)–
O, (μ-O) 2 Cu(II)Cu(III), and (μ-O)(μ-OH)Cu(II)Cu(III) active sites of pMMO. By
looking at the coordination environment of a copper cation in the QM/MM optimized active sites [114] as well as the X-ray crystal structure of pMMO [10], model
complexes were set for DFT calculations to search the methane to methanol conversion. The first model is the Cu(III)–O species coordinated by two imidazole and
T. Yumura et al.
rebound trajectories, Friesner et al. estimated 69–84% retention of stereochemistry
of for the ethane hydroxylation on MMO [80] being close to the corresponding
experimental value (72%) [40].
Furthermore, Friesner group used QM/MM calculations to incorporate the entire
protein environment of sMMO to improve potential energy surfaces of the hydroxylation of several substrates (CH 4 , C 2 H 6 , CH 3 OH, CH 3 CN, and CH 3 NO 2 ) [87].
The QM/MM calculations investigated the energetics of the initial binding of the
substrate into the enzyme, as well as that of the sequent C–H bond dissociation. In
these calculations, van der Waals and electrostatic interactions between the protein
and a substrate can be directly accounted to accurately calculate the activation barriers
for both steps. The QM/MM studies found that the energy barrier for the substrate
diffusion is not negligible compared with that in the C–H bond activation. As a result,
Friesner et al. suggested that the energetic information in the catalytic reactions by
MMOH Q , including the substrate binding and the methane activation, is necessary
to understand the experimental findings on kinetics in the hydroxylation reactions.
6 Mechanism for Methane Hydroxylation on pMMO
After obtaining the X-ray crystal structure of the particulate form of MMO (pMMO)
in 2.8 Å resolution [10], Shiota and Yoshizawa performed DFT and QM/MM calculations to elucidate its catalytic function for the methane hydroxylation [114–117].
Taking the crystal structure of pMMO into account, mononuclear and dinuclear
copper species, located within the soluble regions of the pmoB subunit, were considered as the active site for the methane to methanol conversion [114]. A reaction
of the mononuclear site with O 2 results in the formation of a Cu(III)–O species in
an endothermic manner, and that of dinuclear site leads to a (μ-O) 2 Cu(II)Cu(III)
species in an exothermic manner. The optimized structures for the two active sites
surrounded by the protein environment were obtained in the QM/MM methods, as
shown in Fig. 13. Furthermore, Shiota and Yoshizawa assumed that a (μ-O)(μOH)Cu(II)Cu(III) species is formed as an active site of pMMO by the H-atom
transfer from Tyr374 residue outside the first coordination sphere to assist the activation of the O–O bond [116]. The formation of the (μ-O)(μ-OH)Cu(II)Cu(III)
active site was proposed by an analogy to the oxidation of tyrosine to dopaquinone
by tyrosinase [122]. On the other hand, Chen et al. proposed a (μ 3 -O)trinuclear
Cu(II)Cu(II)Cu(III) active site for the methane hydroxylation, which proceeds via a
concerted oxo-insertion manner [123, 124]. However, such a trinuclear copper site
was not observed in the X-ray crystal structure of pMMO [10].
Here, let us focus on mechanisms for the methane hydroxylation by the Cu(III)–
O, (μ-O) 2 Cu(II)Cu(III), and (μ-O)(μ-OH)Cu(II)Cu(III) active sites of pMMO. By
looking at the coordination environment of a copper cation in the QM/MM optimized active sites [114] as well as the X-ray crystal structure of pMMO [10], model
complexes were set for DFT calculations to search the methane to methanol conversion. The first model is the Cu(III)–O species coordinated by two imidazole and