Pitt Researchers Genetically Engineer Protein to Measure and Display Glucose Levels

PITTSBURGH-—Two University of Pittsburgh researchers have created a protein that can not only measure glucose concentrations but also transmit the information about those levels, as will be reported in the July 15 edition of Analytical Chemistry.

The research could lead to a safer, less invasive procedure for diabetics to monitor their blood sugar levels than the finger-prick method currently in use by more than 100 million diabetic patients worldwide. One application of the research would be to embed the protein in a tiny porous capsule implanted just below the patient's skin. The patient would be able to monitor the glucose level with an optical device that would "read" the color of the protein.

Research Assistant Professor Kaiming Ye and Distinguished Professor of Engineering Jerome Schultz, both in the Department of Bioengineering in Pitt's School of Engineering, genetically engineered a glucose indicator protein—a "living biosensor"—by fusing green fluorescent proteins from a genus of jellyfish to both ends of a glucose binding protein from the bacteria E. coli.

"Our work is on the front line of developing new generation of biosensors called living biosensors that can be used to sense intracellular molecules by microscopy, in real time," said Ye. "This provides a new technique to study the causes of many diseases related to cell metabolism, such as diabetes."

"Typically, biosensors are composed of two elements: a component like an enzyme or antibody that recognizes the presence of another material like glucose and a transducing component that gives a signal that can be measured, such as a colored product from an enzyme reaction or fluorescent molecule," said Schultz, who is director of Pitt's Center for Biotechnology and Bioengineering.

"In this research, we have genetically engineered a hybrid protein that combines both functions: recognition and transduction. Thus, when this protein is exposed to glucose, the amount of fluorescence changes that can be easily picked up by a photocell, like those in a digital camera, providing a continuous method of measuring glucose in body fluids," Schultz explained.

Since the molecule is entirely a protein, with no synthetic chemical dyes involved, it can be produced directly within, and by, living cells.

Thus, by placing the gene for this protein into cells in which knowing the glucose concentration is important, scientists will be able to measure the fluorescence coming from these cells under different conditions, e.g. in the pancreas when exposed to insulin, or in contracting heart cells, or in the brain to understand how cells are involved in different functions.

The protein has applications beyond simply measuring glucose, according to Schultz.

It can be used to understand the metabolism and control in different cell types in various research studies with animals.

Also, this type of protein sensor may become a powerful new tool to understand behavior and the effect of drugs on normal and abnormal cells.

The researchers have tested the protein in vitro in mammalian cells and have made a prototype biosensor with the protein. Schultz said he believes that the technology is about a year away from in vivo testing.

One of the technical issues yet to be overcome is that the protein is not sensitive to glucose changes in the range of glucose concentrations normally found in blood.

"Our protein works at much lower glucose concentrations than those found in blood," said Schultz. "So we are working to 'dumb down' the response of the protein by further genetic engineering."

The patent of the technology is pending.

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