Scientists in Japan are using cells as protein factories.

In protein semisynthesis, a target, modified protein is made by combining a synthetic molecule with a protein. The method can be used to incorporate artificial groups, such as labels, into a protein and so provide a way to study the protein's structure and function. However, protein semisynthesis can be low-yielding and requires high concentrations of reactive components. Now Teruyuki Nagamune and colleagues at the University of Tokyo have improved the efficiency of the process.

Protein semisynthesis often uses a split intein, a section of a protein that can excise itself and reattach the remaining portions - the exteins - to give a newly active protein called the splicing product. Nagamune's group focused on a bacterial intein that splits into two fragments. They fused one fragment to a reductase enzyme, which acts as a receptor, and the other, smaller, fragment to a ligand, in this case a reductase inhibitor. As the inhibitor binds to the enzyme, the two fragments are brought together and their proximity leads to an improved yield of the splicing product.

As a ligand (pink) binds to its receptor, two protein fragments are brought together for improved protein yield

The Japanese researchers adapted the concept in cells to make splicing products of synthetic inteins. They used cells that expressed the fragment bound to the reductase receptor and then incubated the cells with the fragment bound to the ligand. Using a fluorescent assay, Nagamune showed that the ligand binds to the receptor as before, but this time the splicing product is made on the cell surface. 'This represents the first demonstration of a semisynthesis of cell surface protein on living cells,' said Shinya Tsukiji, a co-worker on Nagamune's team.

Nagamune's system works at low concentrations and, since the ligand-attached fragment is short it can be made synthetically and a variety of functionalities can be incorporated easily.

Philip Cole, an expert in protein semisynthesis at Johns Hopkins University, in Baltimore, US, views the work as an elegant merger of small molecule and protein chemical approaches to improving protein semisynthesis. 'The Holy Grail is in vivo protein semisynthesis, which no method works particularly well with, said Cole. 'This approach has the chance to be powerful here.'

In the future, Tsukiji proposes to adopt this approach to incorporate a number of chemical probes such as fluorescent dyes and phosphorylated amino acids into target proteins. '[The system] will be invaluable in cell biology research and biotechnology,' said Tsukiji.

Kathleen Too