VHH CDR-H3 conformation is determined by VH germline usage.

Area of Science:

• Immunology
• Structural Biology
• Biochemistry

Background:

• Single antigen-binding domains, VHHs (nanobodies), derived from camelid heavy-chain antibodies, are crucial for diagnostics, research, and therapeutics.
• VHH structural analysis reveals two main clusters based on CDR-H3 loop conformation: extended and kinked.

Purpose of the Study:

• To classify VHH structures based on CDR-H3 conformation.
• To investigate the relationship between VHH CDR-H3 conformation, germline origin, and antigen-binding properties.
• To understand the impact of germline usage on immune repertoire shaping.

Main Methods:

• Structural analysis of variable domains from llama and alpaca VHHs.
• Classification of VHHs into structural clusters based on CDR-H3 loop conformation.
• Correlation analysis between CDR-H3 conformation, germline gene usage (IGHV3-3, IGHV3S53), and V(D)J recombination patterns.

Main Results:

• VHHs are categorized into two main structural groups: those with extended CDR-H3 loops and those with kinked CDR-H3 loops.
• Extended loops protrude into solvent, while kinked loops fold back onto framework regions, exhibiting distinct properties.
• CDR-H3 conformation strongly correlates with germline origin, with IGHV3-3 derived antibodies showing kinked conformations and IGHV3S53 derived antibodies showing extended conformations.
• No bias from V(D)J recombination was observed, suggesting selection processes influence this correlation.

Conclusions:

• VHH CDR-H3 conformation is significantly influenced by germline gene usage, not V(D)J recombination.
• This germline-driven structural bias impacts antigen interaction and shapes the antibody immune repertoire.
• Findings enhance understanding of how antibody framework properties dictate immune repertoire diversity and function.