Scheme 1. Fabrication of Azido- and Alkynyl-MNPs
diagnosis.9 The success of applying MNPs to biology relies
Because of its high degree of efficiency, complete
specificity, and compatibility with water, the Cu(I)-catalyzed
azide-alkyne cycloaddition18 could potentially be applied to
both in vitro and in vivo19 systems, including drug discov-
ery,20 protein conjugation,17,21 proteomics research,22 virus23
and bacterial24 surface modification, and fabrication of
DNA25 and sugar microarrays.26 Thus far the use of Cu(I)-
catalyzed cycloaddition to functionalize an MNP surface
remains unexplored.27 In this report, we describe the im-
mobilization of diverse molecules onto the MNP surface by
Cu(I)-catalyzed 1,3-dipolar cycloaddition.
on the efficient fabrication of target probes on the particle
surface. Amide bond formation has been the most utilized
method to immobilize probes on the MNP surface.10 Alter-
natively, Shiff’s base formation11 and nucleophilic addition
on epoxide12 also have been commonly used to conjugate
solid materials with probe molecules. However, the applica-
tion of the aforementioned methods in protein conjugation
with a solid support results in random covalent bond
formation that may reduce the protein activity.13 While the
specific orientation of an immobilized target protein on a
solid surface can be achieved by noncovalent interactions,
such as biotin- streptavidin14 and His tag-Ni2+ interac-
tions,15 the immobilized protein may dissociate from the
surface during long-term storage.16 To address this technical
issue, we recently showed that Cu(I)-catalyzed alkyne-azide
[2 + 3] cycloaddition17 is an efficient and site-specific
orthogonal reaction that covalently immobilizes target protein
onto a glass surface. The site-specific immobilized protein
showed higher ligand-binding activity when compared with
protein immobilized by random amide bond formation.
To investigate the reactivity difference between soluble
or immobilized azide and alkyne, the azido and alkynyl
functional groups, respectively, were assembled onto the
MNP (silica oxide-coated Fe3O4).9 The N-hydroxysuccin-
imide (OSu)-activated MNP was incubated with 10 mM
aqueous 3-azidopropanylamine and monopropargylamine,
respectively, for 12 h at 4 °C. The MNP was then capped
with Ac2O/pyridine (1/1) for 3 h followed by ethanolamine
(100 mM) for 1 h to yield the azido- and alkynyl-modified
MNPs (1 and 2), as shown in Scheme 1. The capping steps
(7) Gupta, A. K.; Gupta, M. Biomaterials 2005, 26, 3995-4021.
(8) Gu, H.; Xu, K.; Xu, C.; Xu, B. Chem. Commun. 2006, 941-949.
(9) (a) Chou, P.-H.; Chen, S.-H.; Liao, H.-K.; Lin, P.-C.; Her, G.-R.;
Lai, A. C.-Y.; Chen, J.-H.; Lin, C.-C.; Chen, Y.-J. Anal. Chem. 2005, 77,
5990-5997. (b) Lin, P.-C.; Chou, P.-H.; Chen, S.-H.; Liao, H.-K.; Wang,
K.-Y.; Chen, Y.-J.; Lin, C.-C. Small 2006, 2, 485-489.
(10) Perez, J. M.; Simeone, F. J.; Saeki, Y.; Josephson, L.; Weissleder,
R. J. Am. Chem. Soc. 2003, 125, 10192-10193.
(18) (a) Rostovsev, V. V.; Green, L. G.; Fokin, V. V.; Sharpless, K. B.
Angew. Chem., Int. Ed. 2002, 41, 2596-2599. (b) Tornφe, C. W.;
Christensen, C.; Meldal, M. J. Org. Chem. 2002, 67, 3057-3064.
(19) Sawa, M.; Hsu, T.-L.; Itoh, T.; Sugiyama, M.; Hanson, S. R.; Vogt,
P. K.; Wong, C.-H. Proc. Natl. Acad. Sci. U.S.A. 2006, 103, 12371-12376.
(20) Krasin˜ski, A.; Radic, Z.; Manetsch, R.; Raushel. J.; Taylor, P.;
Sharpless, K. B.; Kolb, H. C. J. Am. Chem. Soc. 2005, 127, 6686-6692.
(21) Natarajan, A.; Du, W.; Xiong, C.-Y.; DeNardo, G. L.; DeNardo, S.
J.; Gervay-Hague, J. Chem. Commun. 2007, 695-697.
(11) Dyai, A.; Loos, K.; Noto, M.; Chang, S. W.; Spagnoli, C.; Shafi,
K. V. P.; Ulman, A.; Cowman, M.; Gross, R. A. J. Am. Chem. Soc. 2003,
125, 1684-1685.
(22) Speers, A. E.; Cravatt, B. F. J. Am. Chem. Soc. 2005, 127, 10018-
10019.
(12) Meador, M. A. B.; Fabrizio, E. F.; Ilhan, F.; Dass, A.; Zhang, G.;
Vassilaras, P.; Johnston, J. C.; Leventis, N. Chem. Mater. 2005, 17, 1085-
1098.
(23) Wang, Q.; Chan, T. R.; Hilgraf, R.; Fokin, V. V.; Sharpless, K. B.;
Finn, M. G. J. Am. Chem. Soc. 2003, 125, 3192-3193.
(24) Link, A. J.; Tirrel, D. A. J. Am. Chem. Soc. 2003, 125, 11164-
11165.
(25) Devaraj, N. K.; Miller, G. P.; Ebina, W.; Kakaradov, B.; Collman,
J. P.; Kool, E. T.; Chidsey, C. E. D. J. Am. Chem. Soc. 2005, 127, 8600-
8601.
(26) Byran, M. C.; Fazio, F.; Lee, H.-K.; Huang, C.-Y.; Chang, A.; Best,
M. D.; Calarese, D. A.; Blixt, O.; Paulson, J. C.; Burton, D.; Wilson, I. A.;
Wong, C.-H. J. Am. Chem. Soc. 2004, 126, 8640-8641.
(27) (a) White, M. A.; Johnson, J. A.; Koberstein, J. T.; Turro, N. J. J.
Am. Chem. Soc. 2006, 128, 11356-11357. (b) Fleming, D. A.; Thode, C.
J.; Williams, M. E. Chem. Mater. 2006, 18, 2327-2334.
(13) Cha, T.; Guo, A.; Zhu, X.-Y. Proteomics 2005, 5, 416-419.
(14) Choi, J.; Lee, J. I.; Lee, Y. B.; Hong, J. H.; Kim, I. S.; Park, Y. K.;
Hur, N. H. Chem. Phys. Lett. 2006, 428, 125-129.
(15) Xu, C.; Xu, K.; Gu, H.; Zheng, R.; Liu, H.; Zhang, X.; Guo, Z.;
Xu, B. J. Am. Chem. Soc. 2004, 126, 9938-9939.
(16) Tomizaki, K.-Y.; Usui, K.; Mihara, H. ChemBioChem 2005, 6, 782-
799.
(17) Lin, P.-C.; Ueng, S.-H.; Tseng, M.-C.; Ko, J.-L.; Huang, K.-T.; Yu,
S.-C.; Adak, A. K.; Chen, Y.-J.; Lin, C.-C. Angew. Chem., Int. Ed. 2006,
45, 4286-4290.
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