Semienzymatic cyclization of disulfide-rich peptides using Sortase A

Area of Science:

• Biochemistry
• Peptide Chemistry
• Drug Development

Background:

• Disulfide-rich cyclic peptides are highly stable and valuable scaffolds for peptide therapeutics.
• Traditional peptide cyclization methods like native chemical ligation have limitations.
• Enzymatic cyclization presents a promising, efficient, and cost-effective alternative.

Purpose of the Study:

• To evaluate the efficacy of Sortase A (SrtA) for head-to-tail cyclization of disulfide-rich peptides.
• To demonstrate the broad applicability of SrtA-mediated cyclization across different peptide structures.
• To establish enzymatic cyclization as a viable strategy for developing therapeutic peptides.

Main Methods:

• Utilized Sortase A (SrtA), a bacterial transpeptidase, for peptide cyclization.
• Employed SrtA's recognition motif (LPXTG) and mechanism involving a thioacyl-linked intermediate.
• Applied the method to cyclize diverse disulfide-rich peptides: kalata B1, α-conotoxin Vc1.1, and sunflower trypsin inhibitor 1.

Main Results:

• Successfully demonstrated SrtA-mediated head-to-tail cyclization of multiple disulfide-rich peptides.
• Showcased the enzyme's effectiveness on peptides ranging from 14 to 29 amino acids with varying disulfide bond numbers.
• Confirmed the formation of cyclic structures with intact disulfide bonds within the peptide backbones.

Conclusions:

• Sortase A is a versatile enzyme capable of efficiently cyclizing various disulfide-rich peptides.
• Enzymatic cyclization via SrtA provides a robust platform for generating stable cyclic peptides for therapeutic applications.
• This study validates enzymatic cyclization as a broadly applicable technology for peptide-based drug discovery.