Scientists have, for the first time, determined the three-dimensional structure of a complete, unmodified G-protein-coupled receptor (GPCR) in its native environment: embedded in a lipid membrane.
The team, led by Stanley Opella, Ph.D. at the University of California, San Diego and Francesca Marassi, Ph.D. at Sanford-Burnham Medical Research Institute, used a technique called NMR spectroscopy to map the arrangement of atoms in one particular GPCR, called CXCR1. Their finding was published by Nature on October 21.
What are GPCRs?
Scientists have long known that cells must have some sort of sensor that allows them to detect external signals like aromas, hormones, and neurotransmitters. Adrenalin, for example, hits the outside of a cell yet manages to trigger changes inside the cell—the “flight or fight” response—without actually entering it. For decades, the link between the outside of a cell and the inside remained unknown—until GPCRs were discovered by Robert J. Lefkowitz, M.D. and Brian K. Kobilka, M.D., a finding for which they were awarded the 2012 Nobel Prize in Chemistry earlier this month.
Here’s how a GPCR works: The receptor is embedded in the cell membrane, with some parts sticking outside the cell and some parts poking inside. When a hormone or other substance binds the receptor on the outside of the cell, the receptor rearranges its shape a little bit. In response, a molecule called G-protein binds to the receptor inside the cell. The bound G-protein then triggers a chain reaction of cellular events that ultimately alter the cell’s behavior. Several laboratories at Sanford-Burnham are currently investigating GPCRs for their roles in cardiovascular function, metabolism, and diabetes.
GPCR drug targets
Because GPCRs are critical for many cellular responses to external signals, they are major targets for drug discovery. At least 30 percent of all therapeutic drugs work by influencing GPCR function in some way. More precise knowledge of the structures of these receptors will allow drug makers to tailor small molecules to better fit specific targets, avoiding collateral hits that can cause detrimental side effects.
Opella, Marassi, and their teams determined the 3D structure of CXCR1, a GPCR that detects the inflammatory signal interleukin 8 on the outside of the cell. The resulting G-protein-triggered cascade inside the cell mobilizes immune and inflammatory responses.
CXCR1 is particularly active in tumor metastasis, making it a major target for anti-cancer drugs. In one example, preclinical studies show that blocking this receptor inhibits undifferentiated stem cells within breast cancer tumors, leading to the death of all tumor cell types and preventing them from seeding new tumors.
“Our finding opens up a whole new set of experiments aimed at designing, rather than blindly screening, drugs that interfere with GPCR activity,” said Marassi, associate professor in Sanford-Burnham’s National Cancer Institute-designated Cancer Center.
Visualizing GPCRs—in 3D
Protein structures are most often determined by reading the diffraction patterns of X-rays beamed at their crystalline form. But crystallizing such large, membrane-embedded molecules is a challenge. Scientists often snip off floppy ends to overcome this problem. However, those changes can alter the shape and activity of critical regions of the protein.
“Our approach was to not touch the protein,” Opella said. “We are working with molecules in their active form.”
“The ability to visualize such a receptor in its intact state and in its natural environment means that we can probe exactly how its different parts move to transmit signals across the cell membrane—we can literally watch it at work,” Marassi added.
Their strategy has revealed a new view of these receptors. Previous reports on GPCRs have all noted seven segments weaving through the membrane. Opella’s group sees an eighth lying on the membrane surface, a trait that at least some other GPCRs must share.
And the loops inside and outside of the cell are structured. “For years people thought the loops were mobile. They’re not,” Opella said. “The signals we get from the loops aren’t any weaker than the other parts of the protein as they would be if they were waving about.”
Opella, Marassi, and colleagues hope their latest finding, along with continuing studies on the changes in the configuration of CXCR1 as it binds to interleukin 8 and drug candidates, will lead to more effective and less harmful cancer treatments.
This study was funded by the National Institutes of Health.
Park, S., Das, B., Casagrande, F., Tian, Y., Nothnagel, H., Chu, M., Kiefer, H., Maier, K., De Angelis, A., Marassi, F., & Opella, S. (2012). Structure of the chemokine receptor CXCR1 in phospholipid bilayers Nature DOI: 10.1038/nature11580