Here, we first report a rapid and
highly selective biocompatible
ligation that proceeds via a strain-promoted prenyl-involved [2, 3]-Ene
rearrangement process. We demonstrate the usefulness of naturally
occurring strain-promoted β-caryophyllene with triazoline (PTAD)/Selectfluor
in the study of tagging molecule-of-interest. Experiments in peptide
(Histone H3 (1–21) and Myhc (614–629)) and protein (BSA,
βLG, and HSP40) models exemplified the utility of the Ene-ligation
for in vivo imaging and tracking.
We report here the selective incorporations of nitroso
species
into a wide range of proteins targeting lysine residue(s). The corresponding
azo functionalities were formed in a highly selective manner with
excellent yields, displaying rather good stability under physiological
conditions. Furthermore, the azodation proceeded smoothly in high
yields on targeted peptides. Fluorescent and/or dual fluorescent labeling
of varied proteins following this protocol have been determined efficiently
and selectively. With this established protocol, we aim to determine
its usage in the evaluation of the interaction of prenylated proteins
with their interacted enzyme(s) via FRET assays. Delightedly, chemically
modified proteins with a 1-pyrenyl fluorophore through 254 nm UV irradiation
and the sequential azodation and click reaction of protein prenyl
functionality, which enable the incorporation of naphthene, indeed
increase the fluorescence energy transferred since we observed significantly
enhanced absorption located at 218 nm in lysed HEK293T cells and a
clearly strengthened greenish fluorescence in living HEK293T cells.
Prenyl functionalities have been widely discovered in
natural products,
nucleic acids, and proteins with significant biological roles in both
healthy and diseased cells. In this work, we develop a series of new
nitroso-based probes for the labeling, enrichment, and regulation
of prenylated RAS protein, which is highly associated with ∼20%
of human cancers and used to be regarded as an “undruggable”
target via a sequential ene-ligation and oxime condensation (SELOC)
process. We found that these nitroso species can rapidly react with
prenyl-containing molecules through ene-ligation and install a molecular
tag for functional applications under physiological conditions. We
first investigated this ligation process on two peptide models and
demonstrated its labeling efficiency on various proteins such as myoglobin,
lysozyme, RNase A, BSA, and HSP40. We further coupled this reactive
platform with proteolysis-targeting chimera technology targeting to
increase its efficiency and accuracy, as well as to expand its application
range. Using the prenylated RAS protein as the model, we demonstrated
that RAS could be efficiently decorated with our nitroso probes, which
further condensate with oxime and rapidly react with a pomalidomide-containing
hydroxylamine probe for protein degradation. As a result, the RAS
protein in both HeLa and A549 cell lines has been determined to be
efficiently degraded both in vitro and in vivo. This is the first
case targeting post-translational modification other than ligand–protein
interaction to degrade and regulate RAS proteins. We envision that
our SELOC strategy will have great potential in studying the fundamental
structures and functions of prenylated biomolecules and developing
new drugs based on these unique cellular pathways.
Cyano group as a versatile functionalized intermediate has been explored for several decades, as it readily transfers to many useful functionalization groups such as amine, amide, acid, etc., which make it possess high popularization and use value in organic synthesis. Reactions involved with element-cyano bond cleavage can provide not only a new cyano group but also a freshly functionalized skeleton in one-pot, consequently making it of high importance. The highlights reviewed herein include H-CN, Si-CN, C-CN, B-CN, Sn-CN, Ge-CN, S-CN, Halo-CN, N-CN, and O-CN bonds cleavages and will summarize progress in such an important research area. This review article will focus on transition metal catalyzed reactions involving element-cyano bond activation.
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