Nanobodies—single-domain antibodies derived from camelids—have become indispensable tools in diagnostics, therapeutics, and targeted drug delivery. However, enhancing their functionality through precise and efficient chemical modification remains a key challenge in the field.

A recent study introduces a chemoenzymatic method for site-specific lysine modification of VHH nanobodies using lysine acylation with conjugating enzymes (LACE), enabling subsequent bioconjugation via potassium acyltrifluoroborate (KAT) ligations. This innovative approach provides a versatile and broadly applicable solution to a long-standing hurdle in nanobody engineering.

How It Works

Researchers expressed VHH nanobodies in Escherichia coli, incorporating a short LACE tag (4 or 11 residues) near the nanobody’s C-terminus. This tag directed site-specific transfer of functional group-bearing peptides to a single lysine residue. The modified lysine served as a handle for rapid and chemoselective KAT ligation, enabling conjugation to a variety of biomolecules—including small molecules, other nanobodies, and full-length antibodies.

Why It Matters

Unlike fusion protein strategies—which can interfere with nanobody folding or activity—this post-expression modification preserves native nanobody structure and function. The mild, aqueous conditions and compatibility with micromolar concentrations make this technique particularly attractive for therapeutic development and multiplexed diagnostics.

This study opens new doors for building customizable nanobody conjugates with enhanced targeting, tracking, or therapeutic payload delivery capabilities.

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Reference: ACS Publications