Environmental Engineering Reference
In-Depth Information
Mechanism 1 is reminiscent of other well characterized enzymatic systems, such
as methionine synthase and methyl-SCoM methyltransferase, where high-energy
methyl-N and methyl-S bonds undergo a nucleophilic substitution by Co(I) yielding
stable methyl-cob(III)alamin intermediates that act as methyl donors in subsequent
reactions [
95
,
96
]. Experimental evidence in support of mechanism 1 includes
the inversion of stereoconfiguration during the reduction of ethyl-CoM [
89
], the
generation of an alkyl-Ni(III) bond during the reaction with the inhibitor
3-bromopropane sulfonate [
84
,
97
,
98
] and related brominated analogs of methyl-
SCoM [
51
,
74
,
82
], and the formation of methyl-Ni(II)F
430
species in the reaction of
free cofactor F
430
with methyl iodide and methyl bromide [
40
]. In addition, the
reaction of MCR with ethyl-CoM exhibits a significantly lower catalytic efficiency
relative to that with methyl-SCoM, thus implying steric hindrance during the S
N
2
substitution.
Based on hybrid DFT calculations, mechanism 1 is suggested to not be thermo-
dynamically feasible because of the high energy barrier (45 kcal/mol) for breaking
the C-S bond of methyl-SCoM relative to that to form a relatively weak methyl-Ni
bond [
92
]. Thus, an alternative mechanism, mechanism 2, was proposed in which a
nucleophilic attack of Ni(I) on methyl-SCoM forms Ni(II)-SCoM and a methyl
radical (instead of methyl-Ni(II)), which is immediately quenched by a hydrogen
atom transfer from CoBSH [
92
,
99
,
100
]. Then, as in mechanism 1, the resulting
thiyl radical (CoBS
•
) reacts with Ni(II)-SCoM to give a disulfide anion radical
(CoBS
•
SCoM
) that transfers an electron to Ni(II) to regenerate Ni(I), thus closing
the reaction cycle. DFT calculations predict the energy barrier for the first and
second step of methanogenesis catalyzed by MCR to be 10 kcal/mol and
20 kcal/mol, respectively, thus making the C-S bond cleavage rate-limiting in
methane formation [
100
]. Recent carbon kinetic isotope effect studies (i.e.,
12
CH
3
-SCoM/
13
CH
3
-SCoM
¼
1.04) also indicate that C-S bond cleavage is rate-
limiting [
91
].
Mechanism 2 includes as a key intermediate a form of the enzyme (at least in
terms of coordination geometry at Ni) that has been well-studied by crystallography
and spectroscopy, i.e., a hexacoordinate Ni(II)-SCoM species [
77
,
101
]. One lim-
itation in mechanism 2 as it was originally proposed is that rotation of a free methyl
radical (
•
CH
3
) would scramble the stereochemistry at carbon. This is inconsistent
with the experimental result that the reaction with ethyl-SCoM leads to net inver-
sion of configuration at carbon. In order to explain a net inversion of configuration
for the released methane molecule, the hydrogen atom transfer from CoBSH has to
be faster than rotation of the methyl radical in the active site. This would likely be
possible only if hydrogen atom abstraction is concerted with formation of a
transient methyl radical as recently suggested [
102
]. A proposed transition state
for the methyl radical intermediate in the concerted methyl radical mechanism is
shown in Figure
4
, with bond distances proposed in the computational studies.
Recent kinetic isotope effect studies are consistent with such a transition state.
A secondary kinetic isotope effect of 1.19 per D in the methyl group of CD
3
-SCoM
was interpreted to indicate a change in geometry of the methyl group from
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