The study of biomolecules in their native environments is a challenging task because of the vast complexity of cellular systems. Technologies developed in the last few years for the selective modification of biological species in living systems have yielded new insights into cellular processes. Key to these new techniques are bioorthogonal chemical reactions, whose components must react rapidly and selectively with each other under physiological conditions in the presence of the plethora of functionality necessary to sustain life. Herein we describe the bioorthogonal chemical reactions developed to date and how they can be used to study biomolecules.

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Bioorthogonal Chemistry: Fishing for Selectivity in a Sea of Functionality

CuI‐Catalyzed Alkyne–Azide “Click” Cycloadditions from a Mechanistic and Synthetic Perspective

Human pharmacokinetics data indicate that i.v. ascorbic acid (ascorbate) in pharmacologic concentrations could have an unanticipated role in cancer treatment. Our goals here were to test whether ascorbate killed cancer cells selectively, and if so, to determine mechanisms, using clinically relevant conditions. Cell death in 10 cancer and 4 normal cell types was measured by using 1-h exposures. Normal cells were unaffected by 20 mM ascorbate, whereas 5 cancer lines had EC50 values of <4 mM, a concentration easily achievable i.v. Human lymphoma cells were studied in detail because of their sensitivity to ascorbate (EC50 of 0.5 mM) and suitability for addressing mechanisms. Extracellular but not intracellular ascorbate mediated cell death, which occurred by apoptosis and pyknosis/necrosis. Cell death was independent of metal chelators and absolutely dependent on H2O2 formation. Cell death from H2O2 added to cells was identical to that found when H2O2 was generated by ascorbate treatment. H2O2 generation was dependent on ascorbate concentration, incubation time, and the presence of 0.5-10% serum, and displayed a linear relationship with ascorbate radical formation. Although ascorbate addition to medium generated H2O2, ascorbate addition to blood generated no detectable H2O2 and only trace detectable ascorbate radical. Taken together, these data indicate that ascorbate at concentrations achieved only by i.v. administration may be a pro-drug for formation of H2O2, and that blood can be a delivery system of the pro-drug to tissues. These findings give plausibility to i.v. ascorbic acid in cancer treatment, and have unexpected implications for treatment of infections where H2O2 may be beneficial.

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Pharmacologic ascorbic acid concentrations selectively kill cancer cells: Action as a pro-drug to deliver hydrogen peroxide to tissues

An Oxygen Scavenging System for Improvement of Dye Stability in Single-Molecule Fluorescence Experiments

Free‐Radical‐Based, Specific Desulfurization of Cysteine: A Powerful Advance in the Synthesis of Polypeptides and Glycopolypeptides

Tris(2-carboxyethyl) phosphine (TCEP) reduces disulfides rapidly and completely in water at pH 4.5. It preferentially reduces more strained disulfides, in accordance with the usual mechanism postulated for reduction of disulfides by phosphines in water. The reagent can be synthesized conveniently in large quantities by acidic hydrolysis of the commercially available tris(2-cyanoethyl) phosphine

James C. Butler

Papers

Selective reduction of disulfides by tris(2-carboxyethyl) phosphine

Selective Reduction of Disulfides by Tris(2‐carboxyethyl)phosphine.