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Selective chemical reactions enacted within a cellular environment can be powerful tools for elucidating biological processes or engineering novel interactions. A chemical transformation that permits the selective formation of covalent adducts among richly functionalized biopolymers within a cellular context is presented. A ligation modeled after the Staudinger reaction forms an amide bond by coupling of an azide and a specifically engineered triarylphosphine. Both reactive partners are abiotic and chemically orthogonal to native cellular components. Azides installed within cell surface glycoconjugates by metabolism of a synthetic azidosugar were reacted with a biotinylated triarylphosphine to produce stable cell-surface adducts. The tremendous selectivity of the transformation should permit its execution within a cell's interior, offering new possibilities for probing intracellular interactions.
Chemoselective ligation reactions designed to modify only one cellular component among all others have provided insight into cellular processes (1). The goal in developing such transformations is to equal the tremendous selectivity of noncovalent recognition events, such as antibody-antigen binding, that direct many normal biological processes and are now powerful experimental tools. In order to achieve this, the two participating functional groups must have finely tuned reactivity so that interference with coexisting functionality is avoided. Ideally, the reactive partners would be abiotic, form a stable adduct under physiological conditions, and recognize only each other while ignoring their cellular surroundings. The demands on selectivity imposed by cells preclude the use of most conventional covalent reactions, and thus far only two have proven utility in a biological environment.
One chemoselective ligation reaction, that between a ketone and an aminooxy or hydrazide group, has enabled us to engineer the composition of cell surfaces (2). We introduced ketones onto cells through unnatural sialic acid biosynthesis. Human cells metabolize the unnatural precursor N-levulinoylmannosamine (compound 2, Fig. 1), a ketone-bearing analog of the native sugar N-acetylmannosamine (compound 1.Fig. 1), to the corresponding keto-sialic acid residues on cell surface glycoconjugates. Chemically orthogonal to native cell surface components, the ketone can then react selectively with externally delivered aminooxy or hydrazide reagents to form stable covalent adducts. Applications of this reaction include the chemical construction of new glycosylation patterns on cells (3), new approaches to tumor cell targeting (4), and novel receptors for facilitating viral-mediated gene transfer (5).
Although useful for cell surface...