Species-Dependent Structural Variations in Single-Domain Antibodies.

Area of Science:

• Biotechnology
• Immunology
• Structural Biology

Background:

• Single-domain antibodies (sdAbs) are derived from camelid heavy-chain antibodies (HCAb).
• sdAbs offer advantages like small size, high stability, and ease of production, making them valuable for biomedical research and therapeutics.
• While numerous sdAb structures are documented, comprehensive analyses of species-dependent structural variations are limited.

Purpose of the Study:

• To evaluate species-dependent structural differences in single-domain antibodies (sdAbs).
• To create a comprehensive, open-access database of sdAbs with known antigen targets.

Main Methods:

• Assembled MO-IISA, an open-access database, by integrating six public resources (iCAN, INDI, SAbDab-nano, sdAb-DB, PLabDab-nano, NbThermo).
• Applied harmonized eligibility criteria for data inclusion.
• Quantified region lengths, amino acid frequency, and conservation/entropy across frameworks (FR1-FR4) for 2053 sdAbs.

Main Results:

• The dataset included 2053 sdAbs from llamas (n=1316), alpacas (n=325), dromedary camels (n=377), and Bactrian camels (n=35).
• Average sdAb length was ~124 amino acids with minor interspecies variations.
• Framework regions (FRs) were highly conserved, while CDR2 and CDR3 exhibited the most inter- and intra-species variability. Lysine enrichment was noted in FR3 and FR2, and cysteines in FR1 and FR3, with non-canonical cysteines more frequent in Bactrian and dromedary sdAbs.

Conclusions:

• Framework regions exhibit species-neutral constraints, while loop regions show species-tuned adaptations.
• These structural features have practical implications for sdAb engineering, species selection, and conjugation strategies.
• The MO-IISA database provides a valuable resource for exploring these species-specific characteristics in sdAbs.