Chemists Show Proteins Behave Differently Inside Cells Than They Do In Water Solutions

"For 40 years, we thought we could learn most everything about proteins by studying them in water, but this work shows we are missing important observations by looking at them just in water or other solutions," said Dr. Gary Pielak, professor of chemistry and lead author of the study. "Our work demonstrates that we need to study them under the conditions they are found in inside the cell."

The research is relevant to medicine because the protein is related to proteins associated with Parkinson's and Alzheimer's diseases and cancer, the scientists say.

"Proteins are the robots of the cell in that they perform countless functions including allowing cells to grow and reproduce," Pielak said. "Almost everything we know about them comes from research done in test tubes in water solutions. But inside cells, where proteins work, there are no dilute solutions because the interior is crowded with proteins, which take up about 40 percent of the volume."

Working under Pielak's supervision, Matthew Dedmon of Gastonia, N.C., used nuclear magnetic resonance (NMR) spectroscopy to examine what effects the crowded environment had on protein shape because the shape of a protein determines its function. The team found that a so-called "intrinsically unstructured" protein, which in water appears to have no fixed structure, shows a definite folded-up shape when inside cells.

Among other things, the experiments involved measuring the proteins with a nucleus of nitrogen known as N-15 and then recording and comparing their NMR spectrum both inside cells and outside cells under artificially crowded conditions.

A report on the findings was scheduled to appear online today (Sept. 13) in the Proceedings of the National Academy of Sciences. A senior when he conducted the experiments last year, Dedmon is now on a National Science Foundation Graduate Rsearch Fellowship at England's University of Cambridge. Other authors are Chetan N. Patel, a doctoral student in chemistry, and Dr. Gregory B. Young, manager of the UNC Biomolecular NMR Facility.

"Scientists had theorized for many years that solutions crowded with molecules would tend to favor molecular shapes that had the smallest surface areas," Pielak said. "In some ways, the explanation of our observation has been around for two centuries – since the time of Le Chatelier," he said. "In the past, however, it has been so difficult to do these experiments that few have even tried." With the new information pouring in from the Human Genome Project and other efforts to identify genes, scientists hope to create models of cellular metabolism, which would advance understanding of health and illness, Pielak said.

"But to make a model of cellular metabolism that would run in a computer, you need to know how tightly these proteins bind to one another and how fast they bind," he said. "All those data so far are from solutions that were mainly water. If there are differences between what we measure in dilute solutions and what occurs in cells, no one will ever be able to model metabolism. That means we need to look more thoroughly at the conditions found inside cells and measure them."

"What impresses me the most about this discovery is the clear demonstration that the environment of proteins in real life situations -- the proximity of billions of other molecules such as lipids, sugars, salts and water, for example -- has a profound influence on their three-dimensional structure," said Dr. Edward T. Samulski, Boshamer and Distinguished professor of chemistry at UNC. "Everyone knows that this three-D structure is essential in biology, but very few investigators have had the courage to look at proteins in the complex soup they actually live in."

The National Science Foundation, the Petroleum Research Fund and the Smallwood Foundation supported the study, said Pielak, also a member of the UNC Lineberger Comprehensive Cancer Center and professor of biochemistry and biophysics at the UNC School of Medicine.