Technique reveals dynamics of bound protein waters

A team of scientists from Case Western Reserve University have used the National Synchrotron Light Source (NSLS) at Brookhaven National Laboratory to develop a technique that pinpoints the location and motion of water molecules bound to proteins.

Using temperature-dependent radiolysis and mass spectroscopy, they are able to identify where water is binding tightly or loosely on the surface of a protein and how it is influencing a protein’s function.

This is one of the most interesting things I’ve done in a number of years

“It’s as if there were a window ledge with a pebble stuck in it, so the window doesn’t shut tightly,” said Mark Chance, director of the Center for Proteomics and Bioinformatics at Case Western University.

“The water is like that pebble. It could be an obstacle to the formation of the protein complex, just like the pebble stops the window closing. Or, it could be in just the right position, as if the window had a small notch carved in just the shape of the pebble. Both of those situations could occur in nature.”

In order to find water that’s tightly bound to a protein, the team chemically activated protein waters with an x-ray beam, which permanently attached the tightly bound waters to the protein.

They then washed out the system and take a measurement of the mass of the protein. A shift in mass tells them where the water is bound tightly.

Measuring the exchange rate of water (16O) versus heavy water (18O) revealed how fast the mass of the peptide changes due to bound water molecules. “

To our surprise, that process was slower than we thought it would be,” Chance said. “The water is more tightly bound than we would have expected, and on a timescale that’s important for the chemistry of protein reactions.”

Chance said this method could give insight into the function of water in enzymes such as hexokinase, which catalyses the first step in oxidisation of food into energy in the body.

The method is also well suited to studying aquaporins - proteins embedded in cell membranes that regulate water flow - or ion channels, which transport ions along with water.

“This is one of the most interesting things I’ve done in a number of years,” Chance said. “People are really excited about the possibilities this opens up to study what water is doing and where it is. And this opportunity to study the structure of water has a bright future at the National Synchrotron Light Source II.”

Chance was recently awarded the largest instrumentation grant from the National Science Foundation for a state-of-the-art wiggler to be used at the beamline he is overseeing at NSLS-II.

The instrument will allow scientist to probe the structures of molecules in vivo and to answer fundamental questions about structural biology.