Finding better painkillers is a high priority for doctors, researchers, and patients.
While opioids, such as morphine, are very effective, they come with unwanted side effects and can be addictive and dangerous.
In order to test new pain-killing drugs, researchers need their corresponding receptors, but producing or collecting large quantities of these receptors has always been challenging. Even more problematic is the fact that the chemistry of these receptors makes it impossible to test them in isolation under typical experimental conditions, such as being suspended in a water-based solution.
An interdisciplinary team of Penn researchers has taken steps to solve those two problems. Led by Renyu Liu, an assistant professor in the Department of Anesthesiology and Critical Care at the Perelman School of Medicine, and Jeffery Saven, an associate professor in the Department of Chemistry in the School of Arts & Sciences, researchers used computer modeling to design a water-soluble variant of the mu opioid receptor that can be grown in large quantities within bacteria.
Starting their study before the structure of this receptor was known, the researchers began with only its gene sequence, meaning they knew the order of the protein’s amino acids but not how they were folded together.
The researchers based their first designs on the structures of similar receptors, and checked them against the structure of the mu opioid receptor in a mouse, which was only first discovered after the Penn team began their study. Encouraged by the degree to which their designs matched, they began to identify the amino acids on the exterior of the structure that prevented the receptor from being suspended in water, as well as some of the amino acids that were potentially toxic to the bacteria they were to be grown in.
Replacing 53 of the protein’s 288 amino acids, the research team introduced the new gene sequence into E. coli bacteria, which were able to produce large quantities of the variant. This enabled the researchers to show its value to future studies by performing functional tests.
“We showed that this water-soluble form of the protein can compete with the native, membrane-based form when binding with antagonists that are fluorescently labeled,” Saven says. “You can watch the fluorescence shift as more of these water-soluble variants are floating around in the solution.”
The team’s computational approach enables further iterations of the variant to be more easily designed, meaning it can be tweaked alongside experimental conditions.
“This is a great product that can do a lot of things,” Liu says. “You can use this variant to look at the structure-function relationship for the receptor, or even potentially use it as a screening tool.”