Optical Control of Translation with a Puromycin-Photoswitch

Abstract

Translation is an elementary cellular process that involves a large number of factors interacting in a concerted fashion with the ribosome. Numerous natural products have emerged that interfere with ribosomal function, such as puromycin, which mimics an aminoacyl tRNA and causes premature chain termination. Here, we introduce a photoswitchable version of puromycin that, in effect, puts translation under optical control. Our compound, termed puroswitch, features a diazocine that allows for reversible and nearly quantitative isomerization and pharmacological modulation. Its synthesis involves a new photoswitchable amino acid building block. Puroswitch shows little activity in the dark and becomes substantially more active and cytotoxic, in a graded fashion, upon irradiation with various wavelengths of visible light. In vitro translation as-says confirm that puroswitch inhibits translation with a mechanism similar to that of puromycin itself. Once incorporated into nascent proteins, puroswitch, reacts with standard puromycin antibodies, which allows for tracking de novo protein synthesis using western blots and immunohistochemistry. As a cell-permeable small molecule, puroswitch can be used for nascent proteome profiling in a variety of cell types, including primary mouse neurons. We envision puroswitch as a useful biochemical tool for the optical control of translation and for monitoring newly synthesized proteins in defined locations and at precise time points.

Triplet-Sensitized Bidirectional Isomerization of Bridged Azobenzene

“…Diazocines also exhibit more efficient photoswitching using longer excitation wavelengths and possess higher quantum yields. [1][2][3][4][5][6][7][8][9][10][11][12][13] As a result, diazocines have found use in emerging fields, such as stimuli-responsive systems, [14][15][16][17][18][19] , photoactuation, 20 and most notably, photopharmacology [21][22][23][24][25][26][27][28] . Unlike conventional azobenzene, whose triplet state properties have been investigated since the 1960s, 29 no studies on the triplet states of diazocine molecules have been published.…”