Site‐Specific Protein Modification through Cu^I‐Catalyzed 1,2,3‐Triazole Formation and Its Implementation in Protein Microarray Fabrication

Abstract

Out of site: A protein expression system was combined with CuI‐catalyzed 1,2,3‐triazole formation to modify a target protein at its C terminus. The immobilized core protein can be modified by diverse small molecules in a site‐specific manner (see picture). Protein conjugation with a diazido linker gives a homodimeric protein, and microarrays can be fabricated by site‐specific covalent binding to functionalized surfaces.

Click Chemistry for Drug Delivery Nanosystems

“…Interestingly, the high catalytic activity of BPDS renders unnecessary the use of a large excess of coupling probes. As previously observed with MNP (109) and microarray slides (110), CuAAC on CPMV containing peripheral azides proceeded more efficiently than with alkynes.…”

“…This faster kinetic of the alkyne counterpart in solution is consistent with the required Cu(I) coordination of the alkyne. A similar preference has been reported when labeling CPMV (16,23) and microarray slides (110). Interestingly, the activity of a MBP covalently immobilized on the MNP at its C terminus by site-specific CuAAC was higher than random amide bond formation, in agreement with the C terminus being distant from the MBP binding site.…”

Technological Development of Antibody Immobilization for Optical Immunoassays: Progress and Prospects

“…The interaction between avidin and biotin has been exploited to conjugate antibodies on a solid surface and has been widely used in the immobilization process (Iversen et al, 2008;Lin et al, 2006Lin et al, , 2010Orth et al, 2003). Biotinylated antibodies react with avidin and other biotin-binding proteins, including streptavidin, neutravidin, tamavidin, and captavidin (Garcia-Aljaro et al, 2009), to generate a biocompatible layer on a surface by exploiting one of the strongest bonds (K d D 10 15 M ¡1 ) (Airenne et al, 1999;Diamandis and Christopoulos, 1991).…”

Specific Enzyme Immobilization Approaches and Their Application with Nanomaterials

“…As such, a purification step of target enzymes may be needed before immobilization [30]. c) Lin et al [65] fused an intein at the C-terminal of maltose binding protein (MBP) and eGFP, and an azido group was introduced into MBP and eGFP by Cys alkyne. Similar method was also used to introduce alkyne groups into eGFP.…”

“…The Cu(I)-catalyzed Huisgen 1,3-dipolar cycloaddition of azides with an alkyne, a well-known as the flagship reaction of the ''click'' chemistry family [62], has been used extensively in modern chemistry from drug discovery to materials science [63], especially for immobilizing azidoor alkyne-containing proteins onto alkyne-or azido-coated surfaces, respectively [64][65][66][67][68][69]. Azido and alkyne ligands are first attached to the target enzyme and the supporting materials, respectively, and then the reactions are carried out between azido and alkyne to form the covalent bond.…”