Labeling of prenylated proteins via Ene-ligation using naturally-occurring citronellol

Prenylations are intensively existed in natural products, post-transcriptional modifications of nucleic acids and post-translational modifications of proteins. We report a [2, 3]-rearrangement Ene-ligation between citronellol and 4-phenyl-3H-1, 2, 4-triazole-3, 5(4H)-dione (PTAD) at ambient temperature. The method displays several advantages, i.e., fast reaction rate, good selectivity, and mild physiological conditions. We have also explored substituent effect of PTAD towards the rate of the Ene-ligation with citronellol. Eventually, the fluorescent probe PTAD-DBCO-Cy5 was subjected to bovine serum albumin (BSA) pre-installed with citronellol and fluorescent tagging has been achieved. The subsequent quantitative analysis of the three types of proteins revealed that the PTAD was rapidly linked to citronellol with 33–50% yields. This method expands the biocompatible ligation toolkit for the study of prenylated biomacromolecules. Thus, this research unveiled the Ene-ligation of PTAD with citronellol displaying great potential in the studies of prenylation transcriptome and proteomics.