Dinitroimidazoles as bifunctional bioconjugation reagents for protein functionalization and peptide macrocyclization

Area of Science:

• Bioconjugation Chemistry
• Protein Engineering
• Synthetic Organic Chemistry

Background:

• Achieving efficient and site-specific protein modification under physiological conditions is a significant challenge in chemical biology.
• Existing bioconjugation methods often face limitations in stability, selectivity, or reaction conditions.

Purpose of the Study:

• To introduce 1,4-dinitroimidazoles as novel bifunctional reagents for protein functionalization and peptide macrocyclization.
• To investigate the chemoselective reactivity of 1,4-dinitroimidazoles with specific amino acid residues under varying conditions.
• To demonstrate the utility of these reagents in creating stable protein conjugates and complex peptide macrocycles.

Main Methods:

• Utilizing 1,4-dinitroimidazoles for chemoselective bioconjugation with cysteine residues under aqueous acidic to neutral conditions via a cine-substitution mechanism.
• Employing 1,4-dinitroimidazoles for reaction with lysine residues in organic solvents with base, proceeding through a ring-opening & ring-close mechanism.
• Synthesizing and applying a bisfunctional 1,4-dinitroimidazole derivative for peptide macrocyclization.

Main Results:

• 1,4-dinitroimidazoles exhibit high efficiency and chemoselectivity for protein bioconjugation with cysteines, forming stable linkages superior to maleimide-thiol conjugates.
• These reagents also react with lysines in organic solvents, providing an alternative conjugation strategy.
• Site-specific protein functionalization with fluorophores and bioactive peptides was successfully demonstrated.
• Facile synthesis of peptide macrocycles, including complex bicyclic peptides, was achieved using a bisfunctional derivative.

Conclusions:

• 1,4-dinitroimidazoles represent a versatile and robust platform for site-specific protein modification and peptide macrocyclization.
• The developed conjugation chemistries offer enhanced stability and control compared to existing methods.
• These reagents significantly advance capabilities in protein engineering and the synthesis of complex peptide architectures.