Sortase A Transpeptidase

Introduction to Sortase A

Sortase A – mediated conjugation is a powerful technique for the precise engineering of enzymatic tools or therapeutic biologics. It enables site-specific attachment of antibodies and proteins to various molecules and materials, including fluorescent labels for imaging and sorting, pharmaceutical compounds, nanoparticles, and even cells for drug delivery.

Mechanism of Sortase A-mediated Conjugation

Sortase A is a transpeptidase from Staphylococcus aureus, responsible for attaching proteins to bacterial cell walls. It specifically recognizes the sequence LPXTG, and cleaves between threonine (T) and glycine (G). The enzyme contains a cysteine residue in its active site, which forms a thioacyl intermediate that reacts with oligo-glycine, covalently linking the protein substrate to the oligo-glycine. As the conjugation reaction occurs under mild conditions, the protein structure and function is well preserved.

Biotechnology employs this specificity to precisely label or modify any protein/antibody of interest either in solution or on cell surfaces, supporting applications in bioengineering, diagnostics, and therapeutics.

To label a protein of interest, an LPXTG sequence is inserted into the protein of interest, and a triglycine-linked label (for example, a fluorophore) as well as Sortase A are added.

Applications of Sortase A – based Protein Labeling

Sortase A – mediated conjugation is a precise tool for protein and antibody engineering. It enables the site-specific protein and antibody attachment to various substrates, such as fluorescent labels for imaging and sorting, compounds, nanoparticles, and cells. The enzymatic approach overcomes the limitations of conventional chemical conjugation methods, which are much harder to control and often compromise protein or antibody functionality.