Facile synthesis of photoactivatable adenosine analogs.

Adenosine and its derivatives are important building blocks of the biological system. They serve as the universal energy currency, amplify intracellular signals for various signal transduction pathways, and can also be used as the co-substrates for enzymatic transformations. The synthesis and regulation of adenosine and its analogs rely on the adenosine binding proteins (ABPs). Dysregulated ABP activity contributes to numerous diseases such as cancer, metabolic disorders, and neurodegenerative diseases. Presently, there is intense interest in targeting ABPs for therapeutic purposes. A large fraction of the human ABP family remains poorly characterized. The need for innovative chemical probes to investigate ABP function in the native biological matrix is apparent. In this study, an adenosine analog, probe 1, with a photoaffinity group and biotin tag was synthesized using concise synthetic strategies. This probe was able to label and capture individual recombinant ABPs with good target selectivity. Probe 1 was also evaluated for its ability to label spiked ABP in complex cell lysates. This chemical probe, together with the labeling and enrichment assay, is of great value to interrogate the biological functions of ABPs and to elucidate their diversity under different physiological conditions.

Development of Second Generation Activity-Based Chemical Probes for Sirtuins.

NAD+ (nicotinamide adenine dinucleotide)-dependent protein deacylases, namely, the sirtuins, are important cell adaptor proteins that alter cell physiology in response to low calorie conditions. They are thought to mediate the beneficial effects of calorie restriction to extend longevity and improve health profiles. Novel chemical probes are highly desired for a better understanding of sirtuin's roles in various biological processes. We developed a group of remarkably simple activity-based chemical probes for the investigation of active sirtuin content in complex native proteomes. These probes harbor a thioacyllysine warhead, a diazirine photoaffinity tag, as well as a terminal alkyne bioorthogonal functional group. Compared to their benzophenone-containing counterparts, these new probes demonstrated improved labeling efficiency and sensitivity, shortened irradiation time, and reduced background signal. They were applied to the labeling of individual recombinant proteins, protein mixtures, and whole cell lysate. These cell permeable small molecule probes also enabled the cellular imaging of sirtuin activity change. Taken together, our study provides new chemical biology tools and future drug discovery strategies for perturbing the activity of different sirtuin isoforms.