Development and structure-guided characterization of a novel ACE2-binding macrocyclic peptide.

Area of Science:

• Biochemistry
• Structural Biology
• Molecular Medicine

Background:

• Angiotensin-converting enzyme 2 (ACE2) is vital for blood pressure and electrolyte balance within the renin-angiotensin-aldosterone system (RAAS).
• ACE2 serves as the primary receptor for SARS-CoV and SARS-CoV-2 spike proteins, mediating viral entry.

Purpose of the Study:

• To identify and characterize novel ACE2-binding molecules.
• To elucidate the structural basis of peptide-ACE2 interactions.
• To explore the potential of ACE2-binding peptides for therapeutic and diagnostic applications.

Main Methods:

• Peptide identification via mRNA display.
• Biochemical characterization of ACE2-peptide binding.
• X-ray crystallography to determine the ACE2-WJL-63 complex structure at 2.2 Å resolution.
• Radiolabeling of WJL-63 with DOTA and evaluation of binding affinity (K_D).

Main Results:

• The macrocyclic peptide WJL-63 was identified and characterized.
• The crystal structure revealed WJL-63 binds to the ACE2 catalytic cleft in an upright mode, distinct from small-molecule inhibitors.
• WJL-63 residues Q4, R7, R11, and R14 are crucial for anchoring the peptide.
• The radiolabeled DOTA-WJL-63 showed a K_D of 90 ± 28 nM in binding assays.
• The binding mode necessitates an open ACE2 conformation.

Conclusions:

• The ACE2-WJL-63 complex structure provides a blueprint for designing compounds that modulate ACE2 conformation.
• WJL-63 serves as a scaffold for developing imaging agents for ACE2 visualization.
• This work opens avenues for developing novel therapeutics and diagnostic tools targeting ACE2.