# Protein-encapsulated fluorogenic probes for the selective detection of endogenous O-GlcNAcase (OGA)

## TL;DR

Researchers developed a new fluorescent probe to selectively detect O-GlcNAcase activity in live cells, improving sensitivity and specificity over existing methods.

## Contribution

A protein-encapsulated fluorogenic probe with enhanced OGA selectivity over HEX through structural modification and HSA encapsulation.

## Key findings

- HSA encapsulation significantly improves OGA detection sensitivity in aqueous solutions and live cells.
- Replacing the acetyl group with a propionyl group increases OGA selectivity over HEX.
- The probe can differentiate cell lines based on endogenous OGA and HEX expression levels.

## Abstract

Ever-increasing evidence confirms the role of O-GlcNAcase (OGA) in mediating cell growth and development as well as pathology and underscores the importance of developing sensitive and selective chemical tools for the study of OGA biology. Here, based on our previously developed protein-encapsulation strategy, we designed and synthesized a series of fluorogenic probes based on resorufin, and assembled their composites with human serum albumin (HSA) for the selective detection of endogenous OGA activity in live cells. We show that host–guest self-assembly with HSA significantly enhances the OGA sensitivity of the probes in aqueous solution and cells. The structure of the complex of a glycoprobe and HSA was resolved by small-angle X-ray scattering. We demonstrate that the replacement of the acetyl group in GlcNAc with a propionyl group results in selectivity for OGA over hexosaminidases (HEX) that unselectively hydrolyze hexosamines. This allows us to differentiate between two cell lines with different endogenous OGA and HEX expression levels, and selectively detect OGA activity in live cells.

A human serum albumin (HSA) encapsulated fluorogenic glycoprobe was developed for the selective detection of O-GlcNAcase (OGA) over hexosaminidase (HEX) in cell lysates and live cells.