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Communication
attack by the carbenoid species. Notably, the beneficial effect of this
substitution for carbene N–H insertion reactivity appeared to be rather
general, as indicated by the high TTNs measured with Mb(L29A) with
other aniline derivatives in addition to those considered above (e.g., 5b:
4990 turnovers; 6b: 6790 turnovers).
In conclusion, this work provides the first demonstration of the
ability of engineered Mb variants to efficiently catalyze carbenoid N–H
insertion reactions with aryl amine substrates. The excellent chemo-
selectivity, high numbers of catalytic turnovers, and broad substrate
scope across variously substituted anilines and aryl amines make
these Mb-derived catalysts a synthetically valuable and competitive
alternative to transition metal-based systems typically employed in
these transformations. These studies pave the way to exploration of
the scope of these Mb-derived catalysts in the context of other classes
of amine substrates as well as other carbene transfer reactions.
This work was supported by the U.S. National Institute of Health
grant GM098628. MS instrumentation was supported by the U.S.
NSF grant CHE-0946653.
Fig. 1 Total turnovers supported by the different Mb variants for the
formation of N–H insertion products 12b (N-methyl aniline + EDA) and
1
5 (aniline + tBDA).
in principle modulated by mutagenesis. Despite the broad substrate
scope and high activity of Mb(H64V,V68A), comparatively lower
yields were observed with N-methyl-aniline as the N–H component
Notes and references
1
(a) M. P. Doyle, M. A. McKervey and T. Ye, Modern catalytic methods
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(Scheme 2). In the interest of determining whether improved
activities on this substrate could be obtained by protein engineering,
we screened a panel of previously prepared Mb variants containing
4
6, 9148; (c) D. Gillingham and N. Fei, Chem. Soc. Rev., 2013, 42, 4918.
2
(a) C. Bolm, A. Kasyan, K. Drauz, K. Gunther and G. Raabe, Angew.
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B. Clapham, G. Koch, J. Zimmermann and K. D. Janda, Org. Lett.,
1
–2 amino acid mutations within the distal cavity of the protein
7
(
Fig. 1). From the screening, both Mb(H64V) and Mb(H64V,L29A)
were found to be considerably more efficient than Mb(H64V,V68A)
toward the conversion of N-methyl-aniline, supporting about nearly
four-fold higher turnovers on this substrate (3910 vs. 1030, Fig. 1, top
panel). Using Mb(H64V,L29A) at 0.2 mol%, 12b could be thus
obtained in higher yields (75%). To examine the performance of
the Mb variants in the presence of a different a-diazoester reagent,
reactions with aniline were repeated with tert-butyl diazoacetate
2004, 6, 4627; (e) A. C. B. Burtoloso and C. R. D. Correia, Tetrahedron
Lett., 2004, 45, 3355; ( f ) J. R. Davies, P. D. Kane and C. J. Moody,
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(
(
c) J. F. Nicoud and H. B. Kagan, Tetrahedron Lett., 1971, 2065;
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(
tBDA) as the carbene source. Whereas this reaction is efficiently
50, 5223; (e) C. F. Garcia, M. A. McKervey and T. Ye, Chem. Commun.,
996, 1465; ( f ) S. Bachmann, D. Fielenbach and K. A. Jorgensen,
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1
catalyzed by Mb(H64V,V68A) (3620 TTN, Fig. 1, bottom panel), even
higher TTN values for the formation of 15 were observed for
Mb(H64V) and Mb(H64V,L29A) (5730 and 7540 TTN, respectively).
Collectively, these experiments illustrate how the substrate reactivity
of these Mb catalysts toward different amine substrates and diazo
reagents can be effectively modulated via modification of the
hemoprotein active site.
An interesting aspect emerging from the studies above concerns the
activity-enhancing effect of the H64V substitution, a trend shared
across other types of Mb-mediated group transfer reactions previously
10, 1529; (k) Z. Hou, J. Wang, P. He, B. Qin, X. Liu, L. Lin and
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4
5
7,8
investigated by our group. Since His64 ‘blocks’ the distal face of the
11
heme to the solvent (Fig. S2 in the ESI†), the observed activity
enhancement could stem from an increased accessibility of the heme
center to the reagents as a result of this mutation. Conversely, the L29A
substitution exhibited a distinctive beneficial effect for N–H insertion
activity (Fig. 1), while the same mutation had essentially no impact on
Mb-catalyzed olefin cyclopropanation, despite a common reactive
intermediate (i.e., heme-bound carbenoid, Scheme 1) is likely involved
in these reactions. As this mutation expands the volume above the
heme iron center (Fe–Leu29(C ) distance B8 Å, Fig. S2 in ESI†), it could
b
6 Z. J. Wang, N. E. Peck, H. Renata and F. H. Arnold, Chem. Sci., 2014,
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5
7
M. Bordeaux, V. Tyagi and R. Fasan, Angew. Chem., Int. Ed., 2014,
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2
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play a role in better accommodating the amine substrate prior or after
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Chem. Commun.