Angewandte Chemie International Edition
10.1002/anie.201708428
COMMUNICATION
MAO-A. Nevertheless, when the MAO-A plasmid-transfected
HepG2 cells were incubated with probe 3, a great fluorescence
enhancement (image c) was observed, and the enhanced
fluorescence can be diminished by about 85% (image d and
Figure 3B) by clorgyline (specific MAO-A inhibitor). This
indicates that the plasmid-transfected HepG2 cells did produce a
high level of MAO-A, as confirmed by western blot analysis
We thank the financial support from the 973 Program (nos.
2015CB932001 and 2015CB856301), the NSF of China (nos.
21535009, 21435007, 21675159 and 21621062), and the
Chinese Academy of Science (XDB14030102). We also thank
Prof. Yalin Tang and Dr. Qian Li for help with docking study.
Keywords: analytical methods • fluorescent probes • new
specific recognition moiety • monoamine oxidase A
(Figure 3C), and probe 3 reacts with intracellular MAO-A only.
a
b
c
d
A
[1]
a) M. D. Berry, A. D. Juorio, I. A. Paterson, Prog. Neurobiol. 1994, 42,
3
75-391; b) M. B. H. Youdim, J. J. Beccafusco, J. Neural. Transm.
image
2005, 112, 519-537; c) M. B. H. Youdim, D. Edmondson, K. F. Tipton,
Nat. Rev. Neurosci. 2006, 7, 295-309.
[
2]
3]
a) C. C. Wang, E. Billett, A. Borchert, H. Kuhn, C. Ufer, Cell. Mol. Life.
Sci. 2013, 70, 599-630; b) J. Saura, Z. Bleuel, J. Ulrich, A.
Mendelowitsch, K. Chen, J. C. Shih, P. Malherbe, M. D. Prada, J. G.
Richards, Neuroscience 1996, 70, 755-774.
DIC
B
C
[
a) D. L. Murphy, S. Lipper, S. Slater, D. S. Ling, Psychopharmacology
MAO-A
1
0
0
.0
.5
.0
1
979, 62, 129-132; b) R. M. Geha, I. Rebrin, K. Chen, J. C. Shih, J. Biol.
0
0
.4 GAPDH
a
b
c
Chem. 2001, 276, 9877-9882; c) D. E. Edmondson, C. Binda, J. Wang,
A. K. Upadhyay, A. Mattevi, Biochemistry 2009, 48, 4220-4230; d) A.
Holt, M. D. Berry, A. A. Boulton, Neurotoxicology 2004, 25, 251-266.
a) R. Mcdermott, D. Tingley, J. Cowden, G. Frazzetto, D. D. P. Johnson,
Proc. Natl. Acad. Sci. USA 2009, 106, 2118-2123; b) K. Yamaguchi, R.
Ueki, H. Nonaka, F. Sugihara, F. Matsuda, S. Sando, J. Am. Chem.
Soc. 2011, 133, 14208-14211; c) S. C. Godar, M. Bortolato, M. P.
Castelli, A. Casti, A. Casu, K. Chen, M. G. Ennas, S. Tambaro, J. C.
Shih, J. Psychiatr. Res. 2014, 56, 1-9.
.2
[
4]
5]
0.0
a
b
c
d
a
b
c
Figure 3. (A) Confocal fluorescence images of HepG2 cells incubated with
probe 3 (5 μM) for 3 h. (a) Unpretreated cells; (b) cells were pre-transfected
with a vector control; (c) cells were pre-transfected with MAO-A plasmid; (d)
cells were pre-transfected with MAO-A plasmid and then pre-treated with
clorgyline (50 μM) for 1 h. The DIC images of the corresponding samples are
shown in the second row. Scale bar, 20 μm. (B) Relative pixel intensity
measurements (n = 3) from the images a-d in panel A (the pixel intensity from
image c is defined as 1.0). (C) Western blot analyses of MAO-A in HepG2
cells from images a-c (glyceraldehyde-3-phosphate dehydrogenase, GAPDH,
was used as a protein standard). The results are the mean SD (n =3).
[
a) X. Li, X. Gao, W. Shi, H. Ma, Chem. Rev. 2014, 114, 590-659; b) X.
Wu, X. Li, W. Shi, Q. Gong, H. Ma, Angew. Chem. Int. Ed. 2016, 55,
1
4728-14732; Angew. Chem. 2016, 128, 14948-14952; c) L. H. Qian, L.
Li, S. Q. Yao, Acc. Chem. Res. 2016, 49, 626-634; d) H. W. Liu, K. Li, X.
X. Hu, L. M. Zhu, Q. M. Rong, Y. C. Liu, X. B. Zhang, J. Hasserodt, F. L.
Qu, W. H. Tan, Angew. Chem. Int. Ed. 2017, 56, 11788-11792; Angew.
Chem. 2017, 129, 11950-11954; e) L. Wang, L.Yuan, X. Zeng, J. J.
Peng, Y.Ni, J. C. Er, W. Xu, B. K. Agrawalla, D. D. Su, B. Kim, Y. T.
Chang, Angew. Chem. Int. Ed. 2016, 55, 1773-1776; Angew. Chem.
In summary, we have proposed a general design strategy
for a fluorescent probe specific for an enzyme by using the
characteristic structure of the enzymatic inhibitor, and this is
successfully demonstrated with MAO-A. By combining the
substituted phenol of clorgyline with propylamine as a new
recognition moiety for MAO-A, resorufin-based probes 1-6 were
prepared. The substituted phenol of clorgyline may serve as a
targeting moiety, whose introduction into the recognition moiety
strengthens the specificity of the probes for MAO-A. As a result,
all of the six probes show higher selectivity for MAO-A than the
control probe without the targeting moiety, among which probe 3
displays the highest specificity. The superior analytical
performance of probe 3 has been demonstrated by imaging SH-
SY5Y and HepG2 cells with different levels of MAO-A, and
further confirmed by the control probe under the same
conditions. Moreover, cell transfection experiments also
validated that probe 3 is capable of distinguishing MAO-A from
MAO-B. Using the proposed strategy, more new MAO-A probes
with different spectroscopic properties may be developed, which
would promote the understanding of MAO-A biofunctions. In
addition, our strategy may be suitable to design specific probes
for other enzymes.
2016, 128, 1805-1808; f) Z. X. Liu, X. Zhou, Y. Miao, Y. Hu, N. Kwon, X.
Wu, J. Yoon, Angew. Chem. Int. Ed. 2017, 56, 5812-5816; Angew.
Chem. 2017, 129, 5906-5910.
[
6]
7]
a) G. Chen, D. J. Yee, N. G. Gubernator, D. Sames, J. Am. Chem. Soc.
2005, 127, 4544-4545; b) W. H. Zhou, M. P. Valley, J. Shultz, E. M.
Hawkins, L. Bernad, T. Good, D. Good, L. T. Riss, D. H. Klaubert, K. V.
Wood, J. Am. Chem. Soc. 2006, 128, 3122-3123; c) A. E. Albers, K. A.
Rawls, C. J. Chang, Chem. Commun. 2007, 4647-4649; d) D. Kim, S.
Sambasivan, H. Nam, K. H. Kim, J. Y. Kim, T. Joo, K. H. Lee, K. T. Kim,
K. H. Ahn, Chem. Commun. 2012, 48, 6833-6835.
[
a) S. Long, L. Chen, Y. Xiang, M. Song, Y. Zheng, Q. Zhu, Chem.
Commun. 2012, 48, 7164-7166; b) L. Li, C. W. Zhang, C. Y. J. Chen, B.
W. Zhu, C . Chai, Q. H. Xu, E. K. Tan, Q. Zhu, K. L. Lim, S. Q. Yao, Nat.
Commun. 2014, 5, 3176; c) L. Li, C. W. Zhang, J. Y. Ge, L. H. Qian, B.
H. Chai, Q. Zhu, J. S. Lee, K. L. Lim, S. Q. Yao, Angew. Chem. Int. Ed.
2015, 54, 10821-10825; Angew. Chem. 2015, 127, 10971-10975.
a) X. Wu, L. Li, W. Shi, Q. Gong, X. Li, H. Ma, Anal. Chem. 2016, 88,
[
[
8]
9]
1
440-1446 ; b) W. Shen, J. Yu, J. Ge, R. Zhang, F. Cheng, X. Li, Y.
Fan, S. Yu, B. Liu, Q. Zhu, ACS Appl. Mater. Interfaces 2016, 7, 928-
35.
9
J. C. Ma, M. Yoshimura, E. Yamashita, A. Nakagawa, A. Ito, T.
Tsukihara, J. Mol. Biol. 2004, 338, 103-114.
[10] a) M. Naoi, W. Maruyama, Y. Akao, H. Yi, Y. Yamaoka, J. Neural.
Trans. Suppl. 2006, 71, 67-77; b) J. C. Shih, K. Chen, Curr. Med. Chem.
2004, 11, 1995-2005.
Acknowledgements
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