Orthogonal Inverse-Electron-Demand Cycloaddition Reactions Controlled by Frontier Molecular Orbital Interactions

Area of Science:

• Chemical Biology
• Organic Chemistry
• Bioconjugation

Background:

• Bioorthogonal chemistry allows reactions within biological systems without interference.
• Simultaneous labeling of multiple biomolecules requires orthogonal reaction pairs.
• Existing methods may lack sufficient orthogonality or broad applicability.

Purpose of the Study:

• To develop novel chemoselective bioorthogonal reactant pairs for simultaneous labeling.
• To exploit frontier molecular orbital interactions for reaction control.
• To demonstrate orthogonal labeling of multiple proteins.

Main Methods:

• Investigated reactivity differences between cyclic dienes, strained alkynes, and isonitriles.
• Utilized transition state frontier molecular orbital interaction energies to predict selectivity.
• Performed simultaneous labeling of two distinct proteins with different fluorophores.

Main Results:

• Established that five-membered cyclic dienes react with strained alkynes but not isonitriles.
• Demonstrated that bulky-substituted tetrazines react with isonitriles, not strained alkynes.
• Successfully achieved orthogonal labeling of two proteins using these distinct reaction pairs.

Conclusions:

• A new strategy for accessing orthogonal click reactions based on orbital interactions was established.
• This approach enables the simultaneous labeling of multiple biomolecules with high selectivity.
• The developed method offers a versatile tool for advanced bioconjugation applications.