Isopeptide Protein Tagging - Isis Project No 4312

From flesh-eating bacteria to protein superglue.

Harnessing a lethal bug

Group A streptococci cause a range of diseases, from the mild “strep throat” to the flesh-eating Necrotising Fasciitis. These bacteria use a number of tricks to enable them to colonise the body and resist the immune system. Thin hairs, called pili, extend from the bacterium, forming strong attachments to human cells. Pili are chains of a protein called the major pilin. These chains spontaneously and irreversibly bind together, making it one of the strongest proteins ever found.

With this system, the peptide can now act as a tag, attached to other proteins in vitro or to proteins on living cells (J.Am.Chem.Soc. 2010. 132, 4526-4527). The tag will also attach to a specially designed binding partner, which itself can be linked to most standard labelling systems or surfaces. The Oxford team refer to the peptide as SpyTag and the protein as SpyCatcher. The glueing action occurs when SpyTag and SpyCatcher meet.

Investigating proteins

Tagging with peptides is routinely used for detection, purification or immobilisation of proteins in biotechnology. Current methods to recognise these protein tags (most often HA, myc, FLAG, 6His varieties) are weak and often require complicated modification.

With this new approach the protein and peptide partners are easy to produce and react irreversibly through an amide bond, simply upon mixing. No other component is required. This lock is stable over time, high temperatures, high forces and with harsh chemical treatment.

Peptide tags are used in immunoassays, nanoassembly, protein purification, cellular imaging and protein arrays. However, the reversibility of binding often compromises sensitivity, life-time and purity.

System highlights

• Composed of the natural 20 amino acids, so it is easy to express.
• Both partners genetically encodable, enabling targeting at a specific place or time.
• Reacts over a wide range of temperatures (4-37^oC).
• No cysteines or requirements for specific buffer components.
• Specificity on human cells.

These features have the potential to overcome current peptide tag limitation.

A family of protein superglues

Proteins containing internal locks are found in Group A streptococci but also in many other bacteria. The Oxford team have recently demonstrated splitting an alternative protein scaffold, where the protein partner is smaller than the pilin and the reaction to the peptide is 1,000-times faster. The same technology could be used to create a family of different protein superglues.

The technology has also been highlighted in the national press.

Patent status

This Oxford invention is the subject of a patent application. Isis is interested in partnering to develop these covalent peptide tags and explore the commercial opportunities. Please contact the Technology Transfer Manager using the link below.

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