Tethered Indoxyl-Glucuronides for Enzymatically Triggered Cross-Linking.

Area of Science:

• Organic Chemistry
• Biochemistry
• Chemical Biology

Background:

• Indoxyl-glucuronides are precursors to indigoid dyes through oxidative dimerization.
• This reaction is catalyzed by β-glucuronidase under physiological conditions.
• Developing novel indoxyl-glucuronides is crucial for expanding their applications.

Purpose of the Study:

• To synthesize novel indoxyl-glucuronide derivatives with conjugatable handles.
• To evaluate the indigoid dye formation of these derivatives using β-glucuronidase.
• To assess the utility of these compounds in bioconjugation chemistry.

Main Methods:

• Synthesis of seven indoxyl-glucuronide target compounds and 22 intermediates.
• Characterization of synthesized compounds.
• Enzymatic assays using β-glucuronidase from different sources and rat liver tritosomes to induce indigoid dye formation.

Main Results:

• Successfully synthesized seven indoxyl-glucuronide derivatives, including four with PEG-based conjugatable handles (azido-PEG, hydroxy-PEG, BCN) and three isomers with PEG-ethynyl groups.
• Demonstrated indigoid dye formation upon treatment with β-glucuronidase.
• Confirmed the chromogenic readout under physiological conditions.

Conclusions:

• The synthesized tethered indoxyl-glucuronides are effective substrates for β-glucuronidase.
• These compounds offer a valuable tool for bioconjugation chemistry.
• The chromogenic readout provides a sensitive method for detecting enzymatic activity and facilitating bioconjugation.