Reagents for metal-free bio-ligation. Ligation of molecules to live cells, tissues, antibodies, proteins, surfaces and magnetic beads
Applications Room-temperature labeling of live cells, tissues,antibodies, surfaces and magnetic proteins without the need for cytotoxic metalcatalysts or of organic solvents. Connection (or ligation) of two fragments to make a largermolecule or structure is often achieved with the help of the so-called"click chemistry". This term is used to describe a set of bimolecularreactions that meet the following criteria: reactions should be wide in scopebut selective; produce high yield of the product, proceed with reasonable rateunder mild conditions; and tolerate broad range of solvents. Among known clickreactions is the reaction of azides with acetylenes. The formation of1,2,3-triazoles in 1,3-dipolar cycloaddition of azides to triple bond is known,but 'because the activation energy of acetylene-azide cycloaddition isrelatively high (ΔG‡ ~26 kcal/mol), the reaction is very slow under ambientconditions. The utility of the reaction of azides with alkynes was expanded bythe discovery of Cu (I) catalysis. 1,3-cycloaddition of azides to terminalacetylenes in the presence of catalytic amounts of cuprous salts is facile atroom temperature in organic or aqueous solutions. The copper-catalyzed versionof the acetylene- azide cycloaddition (a.k.a. azide click reaction) found abroad range of applications from microelectronics to virus labeling, to drugdevelopment. However, the use of cytotoxic Cu (I) catalyst has largelyprecluded application of this click reaction in living systems.Catalyst-free 1,3-dipolar cycloaddition of azides tocyclooctynes has made possible a bio-compatible version of the azide clickreaction. The triple bond incorporated in an eight-membered ring is apparentlyalready bent into the transition state-resembling geometry, thus reducing theactivation barrier. New methods for ligating fragments to make a largermolecule or structure and allow the use of these structures for in vivo studiesare needed.
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