Advancing Nitrile-Aminothiol Strategy for Dual and Sequential Bioconjugation

Area of Science:

• Chemical Biology
• Bioconjugation Chemistry
• Biomolecular Engineering

Background:

• Nitrile-aminothiol conjugation (NATC) is a biocompatible ligation technique known for its high chemo-selectivity.
• Developing novel conjugation strategies is crucial for advancing chemical biology and creating complex bioconjugates.

Purpose of the Study:

• To investigate the reactivity and substrate scope of NAT conjugation chemistry.
• To develop a novel pH-dependent orthogonal NATC for chemical biology applications.
• To demonstrate the utility of NATC for sequential and dual conjugation of biomolecules.

Main Methods:

• Studied reaction kinetics of heteroaromatic nitrile and aminothiol pairings.
• Developed and applied pH-dependent orthogonal NATC for sequential and dual conjugation.
• Synthesized fluorescently labeled antimicrobial peptide-oligonucleotide complex as a dual conjugate.

Main Results:

• Identified an optimal, pH-dependent bioorthogonal pairing between heteroaromatic nitrile and aminothiol groups.
• Achieved rapid (≈1 hour) and high-yield (>90%) conjugation strategies.
• Successfully applied NATC to diverse biomolecules including oligonucleotides, chelates, small molecules, and peptides.
• Demonstrated dual conjugate imaging in live cells using a synthesized peptide-oligonucleotide complex.

Conclusions:

• The developed sequential NATC approach provides a versatile tool for chemical biology.
• NATC exhibits exceptional adaptability for rapidly creating structurally complex bioconjugates.
• This chemistry holds significant potential for fundamental and translational biomedical research.