Journal of the American Chemical Society
detected any dithionite trapped products with the native subꢀ
Page 4 of 5
ACKNOWLEDGMENT
1
2
3
4
strate 1 and various other substrate analogues reported in this
We would like to acknowledge Dr. Howard Williams, Department
of Chemistry, Texas A&M University, for performing cryoprobe
NMR of compounds 22 and 25.
and previous work12.
5
ABBREVIATIONS
6
7
A, adenine; Adoꢀ, 5′ꢀdeoxyadenosyl radical; AdoꢀH, 5′ꢀ
deoxyadenosine;
Ad,
adenosine;
NBDꢀF,
4ꢀfluoroꢀ7ꢀ
8
nitrobenzofurazan.
9
REFERENCES
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
Figure 10: NosLꢀcatalyzed generation of the sulfuramidite
(31) with Lꢀtryptophanamide (27).
1.
Shen, B.; Liu, W., Nat Chem Biol 2011, 7 (3), 154ꢀ160.
2. Mehta, A. P.; Abdelwahed, S. H.; Mahanta, N.;
Fedoseyenko, D.; Philmus, B.; Cooper, L. E.; Liu, Y.;
Jhulki, I.; Ealick, S. E.; Begley, T. P., Journal of Biological
Chemistry 2015, 290 (7), 3980ꢀ3986.
Zhang, Q.; Li, Y.; Chen, D.; Yu, Y.; Duan, L.;
In conclusion, we have demonstrated that Nαꢀ
alkylated tryptophan analogs are substrates for NosL and that
the alkyl group (R) can alter the regiochemistry of the initial
hydrogen atom abstraction from the amino group (R=H) to the
alkyl group (R=Methyl) or to the αꢀcarbon (R=cyclopropyl).
We have also demonstrated that replacing the substrate amine
with a ketone or an alkene changes the 5′ꢀdeoxyadenosyl radiꢀ
cal chemistry from hydrogen atom abstraction to double bond
addition and we propose that this strategy may be useful for
the design of bisubstrate inhibitors of radical SAM enzymes.
We have also demonstrated that the 5′ꢀdeoxyadenosyl radical
can add to one of the sulfurs in the [4Feꢀ4S] cluster and that
dithionite can be used to trap radicals at the NosL active site
with substrate analogues suggesting that this reagent has poꢀ
tential for the trapping of intermediates at the active sites of
radical SAM enzymes. Preliminary experiments with MoaA,
BzaF, MqnC, and MqnE suggest that many of these findings
will have some generality in radical SAM enzymology.
3.
Gerlt, J. A.; Bouvier, J. T.; Davidson, D. B.;
Imker, H. J.; Sadkhin, B.; Slater, D. R.; Whalen, K. L.,
Biochimica et Biophysica Acta (BBA)-Proteins and
Proteomics 2015, 1854 (8), 1019ꢀ1037.
4.
Zhang, Y.; Zhu, X.; Torelli, A. T.; Lee, M.;
Dzikovski, B.; Koralewski, R. M.; Wang, E.; Freed, J.;
Krebs, C.; Ealick, S. E., Nature 2010, 465 (7300), 891ꢀ896.
5.
Demick, J. M.; Lanzilotta, W. N., Biochemistry
2011, 50 (4), 440ꢀ442.
6.
Broderick, J. B.; Duffus, B. R.; Duschene, K. S.;
Shepard, E. M., Chemical Reviews 2014, 114 (8), 4229ꢀ
4317.
7.
Horitani, M.; Byer, A. S.; Shisler, K. A.; Chandra,
T.; Broderick, J. B.; Hoffman, B. M., Journal of the
American Chemical Society 2015, 137 (22), 7111ꢀ7121.
ASSOCIATED CONTENT
Supporting Information
8.
Nicolet, Y.; Zeppieri, L.; Amara, P.; Fontecilla-
Camps, J. C., Angewandte Chemie International Edition
2014, 53 (44), 11840ꢀ11844.
Detailed procedures for substrate synthesis, enzymatic reaction
conditions, HPLC and LCꢀMS chromatograms of all enzymatic
reaction products, and NMR spectra of synthesized compounds
are provided. The Supporting Information is available free of
charge on the ACS Publications website.
9.
Ji, X.; Li, Y.; Jia, Y.; Ding, W.; Zhang, Q.,
Angewandte Chemie International Edition 2016, 55 (10),
3334ꢀ3337.
10.
Sicoli, G.; Mouesca, J.ꢀM.; Zeppieri, L.; Amara,
P.; Martin, L.; Barra, A. L.; FontecillaꢀCamps, J. C.;
Gambarelli, S.; Nicolet, Y., Science 2016, 351 (6279),
1320ꢀ1323.
AUTHOR INFORMATION
Corresponding Author
11.
Chemie International Edition 2015, 54 (31), 9021ꢀ9024.
12. Bhandari, D. M.; Xu, H.; Nicolet, Y.; Fontecillaꢀ
Ji, X.; Li, Y.; Ding, W.; Zhang, Q., Angewandte
*Department of Chemistry, Texas A&M University, Colꢀ
lege Station, Texas 77842.
Phone: (979) 862ꢀ4091.
Camps, J. C.; Begley, T. P., Biochemistry 2015, 54 (31),
4767ꢀ4769.
Eꢀmail: begley@chem.tamu.edu
13.
Imai, K.; Watanabe, Y., Analytica Chimica Acta
1981, 130 (2), 377ꢀ383.
14.
Mahanta, N.; Fedoseyenko, D.; Dairi, T.; Begley,
Funding Sources
T. P., Journal of the American Chemical Society 2013, 135
(41), 15318ꢀ15321.
This research was supported by the Robert A. Welch Foundation
(Aꢀ0034)
15.
Mehta, A. P.; Hanes, J. W.; Abdelwahed, S. H.;
Hilmey, D. G.; Hänzelmann, P.; Begley, T. P.,
Biochemistry 2013, 52 (7), 1134ꢀ1136.
Note
16.
Mehta, A. P.; Abdelwahed, S. H.; Fenwick, M. K.;
Hazra, A. B.; Taga, M. E.; Zhang, Y.; Ealick, S. E.; Begley,
T. P., Journal of the American Chemical Society 2015, 137
(33), 10444ꢀ10447.
While this paper was under review, Professor Zhang also
reported the addition of the adenosyl radical to methylene
analogue of tryptophan.25
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