A speed-dating service for proteins
At the MDC, Erich Wanker is a good person to see for advice on protein relationships. His lab analyzes interactions between these molecules using an impressive array of high-throughput technologies, capable of carrying out the molecular equivalent of speed dating. Tens of thousands of molecules can be rapidly screened against as many potential partners. Protein-protein binding is central to all cellular processes; finding new associations and demonstrating direct interactions are crucial pieces of evidence in determining molecular functions.
Two main technologies are used in the “Interactomics” platform established by Erich’s lab. The first is based on a “classic” approach called a yeast-two-hybrid (Y2H) screen, which was developed about 25 years ago. The method uses yeast cells containing baits – binding domains from proteins that have been attached to a reporter system. The bait is introduced into a strain of yeast along with a “prey” molecule that might interact with it; cells in which this happens change color, so that they can be detected, and the interaction allows the cells to grow on a special medium.
“This is a very fast, efficient method that can be used to validate entire networks of protein-protein interactions as well as to answer more specific questions about smaller networks,” Erich says. “The costs are relatively low and the entire system is automated and designed for high throughput.”
The platform has added two enhancements of the traditional Y2H system: MYTH, which enables scientists to study proteins bound to membranes or molecules they interact with, and CytoY2H, which detects interactions between transcriptionally active proteins. These methods can’t yet be used to study protein-protein interactions in a high-throughput manner, Erich says, but they give a closer look at an aspect of molecules that has been difficult to observe.
The lab has developed another technology called the DULIP (dual luminescence-based co-immunoprecipitation) assay, which is based on similar principles but operates in mammalian cells and can be used to map their interactomes. Robotics innovations developed by the group have turned the assay into a high-throughput method with advantages over traditional co-immunoprecipitation and Western blotting assays, in terms of speed and sensitivity. “This allows us to precisely control each step in the screening procedure,” Erich says. “We obtain much better results than with the classical assays; it allows us to quantify interactions as well as to assess the effects of point mutations on affinities.”
For years Erich’s lab has mainly used the equipment to map the protein interactions responsible for processes and pathologies surrounding neurons. Their work has helped unravel some of the networks of molecules that promote the development of Huntington’s disease, the behavior of amyloid fibers in Alzheimer’s, and other neurodegenerative conditions. While defects in single molecules may trigger pathological processes, many more proteins are usually involved in bringing diseases to their full-blown forms. Clarifying details of their interactions has helped Erich and his colleagues identify weak points in the structures of fibers and aggregations that make good targets for inhibitors.
Erich calls his approach to these issues “Neuroproteomics”, and the technology platforms developed by his lab have central roles to play. A couple of years ago, interest from other groups led the MDC to support the technology as a core facility under the title “Interactomics.”
