Engineering protein biosensors for the point of care

About 1 in every 13 000 people is born with a condition called phenylketonuria. Their bodies can’t get rid of phenylalanine, an amino acid generated when protein is broken down in the digestive tract, and so it can build rapidly to toxic levels. Although the disorder is incurable, infants who are diagnosed and treated immediately can grow up to live almost normal lives, as long as they carefully monitor their phenylalanine levels and alter their diet accordingly. But today’s tools for measuring concentrations of phenylalanine, and of metabolites that are altered in other diseases, are not fast enough for use at the point of care.

Now a group led by Kai Johnsson of the Max Planck Institute for Medical Research in Heidelberg, Germany, has engineered a semisynthetic protein that can simply, rapidly, and accurately sense quantities of phenylalanine and other molecules in body fluids. The researchers use bioluminescence resonance energy transfer (BRET), which involves measuring the energy transferred between two light-sensitive molecules—a fluorescent ligand and a luminescent protein called luciferase. To create the molecular sensor, a receptor protein is fused to luciferase to form a protein chimera that, in turn, is linked to a fluorescent ligand that binds to the receptor only when the dinucleotide NADPH is present. As shown on the right side of the figure, the binding brings the ligand and the fluorophore close enough to the luciferase so that the color of the emitted light changes from blue to red; the color of the emitted light indicates the amount of NADPH.

The ability of the molecular sensor to help those with phenylketonuria derives from the fact that NADPH is produced when phenylalanine is oxidized by NADP⁺. The researchers demonstrated the effectiveness by mixing dilute blood samples with a buffer containing NADP⁺ and the molecular sensor. The sample was then dropped onto paper, where fluorescence was measured and the phenylalanine concentration determined using a digital camera. The concentration measurements were as precise as those determined by standard methods. The new technique, however, is fast and simple enough to measure phenylalanine—or any clinically relevant molecule that is oxidized by NADP⁺—close to the point of care. The hope is that a test can be developed for use in a family doctor’s office. (Q. Yu et al., Science 361, 1122, 2018 .)