Synthetic probes can track activity of proteins in cells.

Synthetic probes can track activity of proteins in cells.

To understand the full complexity of protein interactions within a cell, scientists are developing a range of new analytical technologies. One of these technologies involves labelling proteins with synthetic probes, such as fluorophores and quantum dots, as a mechanism to track the activity of individual protein species within a cell.

So far, this technology has mostly been developed for analysing intracellular proteins. However, chemists from the &0x00C9;cole Poly-technique F?d?rale de Lausanne, Switzerland, and the Institute of Toxicology and Genetics at the Forschungszentrum Karlsruhe, Germany, have now developed the technology for labelling proteins on the surface of cells.

This new technology is based on a natural biological process in which a protein known as acyl carrier protein (ACP) is modified by the enzyme phosphopantetheine transferase (PPTase). Specifically, this process involves PPTase transferring the compound 4’-phosphopantetheine from coenzyme A (CoA) to a serine residue of ACP.

In previous research, scientists have shown that if synthetic compounds are attached to the phosphopanatheine section of CoA then they are also transferred to ACP. The Swiss and German team realised that if it developed cells in which ACP was attached to a specific cell surface protein, then this offered a way to label those proteins with synthetic probes.

To test their idea, the researchers developed a strain of the yeast Saccharomyces cerevisiae, in which ACP was expressed on the cell surface protein a-agglutinin receptor Aga2p. They then incubated these yeast cells with versions of CoA with the fluorescent dye Cy3 attached. In a separate experiment, they also incubated the yeast cells with CoA combined with the protein biotin and subsequently incubated the cells with CdSe quantum dots conjugated with streptavidin, which binds to biotin. In both cases, the compounds attached to CoA were successfully transferred to the ACP on the surface of the yeast cells, causing them to fluoresce.

Similar experiments using mammalian cells were equally successful.

As well as providing a means to investigate the behaviour of cell surface proteins, the labelling technology could become an important tool in the relatively new field of cell surface engineering.

Jon Evans