Copper assisted sequence-specific chemical protein conjugation at a single backbone amide

Area of Science:

• Biotechnology
• Chemical Biology
• Protein Engineering

Background:

• Site-specific protein functionalization is crucial for biotechnology but challenging due to difficulties in differentiating single sites on large proteins.
• Existing methods often lack the required precision, speed, or applicability to native proteins.

Purpose of the Study:

• To develop a novel, rapid, and site-specific method for protein functionalization.
• To demonstrate the versatility and efficacy of the developed method in creating modified recombinant proteins.

Main Methods:

• Development of the Copper Assisted Sequence-specific conjugation Tag (CAST) strategy utilizing metal binding targeting.
• Application of CAST for rapid (second order rate 8.1 M⁻¹s⁻¹) and accurate chemical modification of protein backbones.
• Preparation of various on-demand modified recombinant proteins, including antibody-drug conjugates.

Main Results:

• CAST enables rapid and highly accurate, site-specific protein backbone chemical modification.
• Demonstrated versatility through the creation of homogeneous antibody-drug conjugates with high plasma stability.
• Achieved potent efficacy of the modified proteins in vitro and in vivo.

Conclusions:

• CAST provides an efficient and quantitative method for site-specific payload attachment on large, native proteins.
• This technology advances protein engineering and opens new avenues for biotechnological applications.