Since the early 1990s, scientists have used various methods to generate portraits of how proteins in the human body communicate and interact with each other. Although we have begun to map the human proteome, uncovering and accurately describing the various interactions between and among proteins in intracellular molecular machines and signaling networks will take considerably more research. By creating comprehensive, “standard” protein–protein interaction (PPI) maps in normally operating cells, pathologists can compare these with the PPI maps of diseased cells to find anomalies that may relate to disease etiology. The hope is that by locating deviations in the interactome of diseased cells, we can formulate potential interventions.
In 2012, Biogen Idec and the National Institutes of Health jointly funded the Human Interactome Project at Harvard Medical School. The project aims to generate a high-quality atlas describing the physical organization of proteins and their interconnections in a common human cell—Human Embryonic Kidney 293 (HEK293). Now, just over a year into the project, the Harvard lab—led by professor of cell biology Steven Gygi—has expressed, purified and catalogued the proteins in roughly 13,000 genes from HEK293 cells in the ORFeome collection.
Because the project uses an immortalized cell line, researchers won’t be able to generate a complete PPI map of a dynamic interactome. The up side of this is that they are able to standardize HEK293 cells to benchmark interactomes across different research groups, and to establish a baseline for a draft map of a complete interactome in a human cell.
Using affinity purification coupled to mass spectrometry (AP-MS), the lab has created a very accurate platform for protein identification. The AP-MS platform also allows the team to measure simultaneous interactions among multiple proteins that form complexes to act in cells. From this MS information, they will produce a baseline interactome map of healthy HEK293 cells. The process is complicated and requires the expression, isolation and analysis of thousands of proteins—not to mention the ruling out of false-positive interactions.
“The lab is discovering thousands and thousands of protein products in a completely unbiased approach,” says Gygi. “We are not targeting specific products. We are simply documenting the proteins we find and working through their interactions one by one.”
Once the baseline for the HEK293 healthy cell line is established, the team will work on other representative cell lines and characterize the interactomes for unhealthy cells in a number of disease states. The goal is to generate PPI models for various disease phenotypes, as well as for healthy cells. Pharmaceutical designers can use variations between the two to identify potential therapeutic targets.
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