Enzymatic deacetylation of a small-molecule probe induces optical changes that enable, in a single-step, unprecedented real-time detection of HDAC activity in samples of purified enzymes and cell lysates.
H218O under the bridge: Recently, the deoxyxylulose phosphate (DXP) pathway was discovered to be a second pathway supplying isoprenoid biosynthetic precursors. One of steps is an IspG‐catalyzed reductive deoxygenation of methylerythritol cyclodiphosphate (MEcPP) to 4‐hydroxyl‐3‐methyl‐2‐(E)‐1‐diphosphate (HMBPP). Using [2‐13C,18O]‐MEcPP, we detected the positional isotopic exchange for the bridging oxygen in MEcPP.
Histone deacetylases regulate the acetylation levels of numerous proteins and play key roles in physiological processes and disease states. In addition to acetyl groups, deacetylases can remove other acyl modifications on lysines, the roles and regulation of which are far less understood. A peptide-based fluorescent probe for single-reagent, real-time detection of deacetylase activity that can be readily adapted for probing broader lysine deacylation, including decrotonylation, is reported. Following cleavage of the lysine modification, the probe undergoes rapid intramolecular imine formation that results in marked optical changes, thus enabling convenient detection of deacylase activity with good statistical Z' factors for both absorption and fluorescence modalities. The peptide-based design offers broader isozyme scope than that of small-molecule analogues, and is suitable for probing both metal- and nicotinamide adenine dinucleotide (NAD )-dependent deacetylases. With an effective sirtuin activity assay in hand, it is demonstrated that iron chelation by Sirtinol, a commonly employed sirtuin inhibitor, results in an enhancement in the inhibitory activity of the compound that may affect its performance in vivo.
The inside cover picture shows that the enzyme IspG in the deoxyxylulose phosphate pathway can mediate a reversible CÀO bond cleavage at an inert position. This hypothesis was supported by the detection of diphosphate ester bridging and terminal oxygen atom exchange.
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