Highly Selective Lysine Acylation in Proteins Using a Lys-His Tag Sequence

Area of Science:

• Biochemistry
• Chemical Biology
• Protein Engineering

Background:

• Chemical modification of proteins is crucial for diverse applications.
• Achieving high selectivity for lysine modification remains a challenge.
• Previous work demonstrated N-terminal His tags for selective Nα-amine acylation.

Purpose of the Study:

• To develop a method for selective acylation of lysine Nε-amines.
• To introduce Lys-His tags for targeted protein modification.
• To demonstrate the versatility and applicability of the Lys-His tag system.

Main Methods:

• Design and synthesis of Lys-His peptide tags (Hisn-Lys-Hism).
• Incorporation of Lys-His tags into proteins at various sites (C-terminus, loops).
• Acylation of tagged proteins using simple acylating agents under mild conditions.

Main Results:

• Lys-His tags facilitate highly selective acylation of the designated Lys Nε-amine.
• Modification occurs under mild conditions with high selectivity over native lysine residues.
• The method was successfully applied to acylate the therapeutic antibody Rituximab, even in mixed protein samples.

Conclusions:

• Lys-His tags offer a flexible and efficient strategy for site-specific protein acylation.
• This method overcomes previous selectivity challenges in protein modification.
• The technology holds significant potential for bioconjugation, antibody modification, and therapeutic development.