Israeli scientists have developed a simple activity assay for an enzyme linked to the HIV virus life cycle.

Protein kinase enzymes regulate many cellular pathways, including metabolism and cell movement. While the enzymes are essential for normal cellular function, abnormal protein kinase activity has been implicated in a number of diseases. So assays for protein kinase activity and kinase inhibitor screening have great potential use in medical science laboratories.

Now, Itamar Willner and co-workers from the Hebrew University of Jerusalem have developed an activity assay for the protein kinase casein kinase II (CK2), a target of some HIV-1 transcription inhibitors.

Since protein kinases work by attaching phosphate groups to proteins, the team took advantage of the charged nature of the phosphate products to use a field effect transistor (FET) in the assay. A FET registers a change in conductivity as charged species attach to a gate on its surface. In Willner's assay a peptide that is recognised by CK2 is attached to the gate and exposure of the FET to CK2 and adenosine 5'-triphosphate (ATP) results in phosphorylation of the peptide. This changes the charge at the gate, indicating CK2 activity.

A field effect transistor (top left) detects protein kinase (red) activity as a change in conductivity.

Most protein kinase activity assays monitor either take-up of radiolabelled ATP or fluorescently labelled antibodies as they bind phosphorylated amino acid residues. These typically have poor specificity for the protein kinase of interest and limited sensitivity. In contrast, Willner's assay is specific for CK2 and is label-free, removing the potential hazard of any radioactive, toxic or carcinogenic markers and cutting sample-processing steps. Furthermore, said Willner, 'the sensor reveals good performance in terms of sensitivity, reusability, versatility and ease of operation.'

The Israeli team aims to extend its assay to analyse other kinases and to test potential kinase-targeting drug candidates. They anticipate that as more protein kinase-controlled bioprocesses are discovered, quantitative assays for these enzymes will be in demand.

Freya Mearns