A New Fluorogenic Small-Molecule Labeling Tool for Surface Diffusion Analysis and Advanced Fluorescence Imaging of β‑Site Amyloid Precursor Protein-Cleaving Enzyme 1 Based on Silicone Rhodamine: SiR-BACE1

β-site APP-cleaving enzyme
1 (BACE1) is a major player in
the pathogenesis of Alzheimer’s disease. Structural and functional
fluorescence microscopy offers a powerful approach to learn about
the physiology and pathophysiology of this protease. Up to now, however,
common labeling techniques require genetic manipulation, use large
antibodies, or are not compatible with live cell imaging. Fluorescent
small molecules that specifically bind to the protein of interest
can overcome these limitations. Herein, we introduce SiR-BACE1, a
conjugate of the BACE1 inhibitor S-39 and SiR647, as a novel fluorogenic,
tag-free, and antibody-free label for BACE1. We present its chemical
development, characterize its photophysical and pharmacologic properties,
and evaluate its behavior in solution, in overexpression systems,
and in native brain tissue. We demonstrate its applicability in confocal,
stimulated emission depletion and dynamic single-molecule microscopy.
The first functional studies with SiR-BACE1 on the surface mobility
of BACE1 revealed a markedly confined diffusion pattern.