New crystallization method to ease study of protein structures

March 10, 2008
New crystallization method to ease study of protein structures
Ribbon representation of NE2398, a protein from the Nitrosomonas europaea bacterium. Dotted lines represent the parts of the protein digested with protease. Blue molecules represent other molecules in the crystal lattice.

Researchers at the Midwest Center for Structural Genomics (MCSG), the Structural Genomics Consortium (SGC) and the Structural Biology Center (SBC) at the U.S. Department of Energy's Argonne National Laboratory have developed a new technique for crystallizing proteins that will ease experimentation into protein structures.

In order to study protein structures, biologists must turn what is essentially a soup of purified protein into crystals that have a consistent and ordered structure. Each protein consists of a chain of amino acid subunits that twists into helices, ribbons and loops. Some proteins have less tidy molecular structures than others; in these, disordered amino acid chains dangle off the protein like split ends.

Crystallizing proteins that contain many of these flexible regions takes much more work and patience than working with more organized ones, said Argonne senior biologist Andrzej Joachimiak, who led the Argonne research effort. "We've tried to find a way to remove the disordered parts using computer modeling, but that's been a challenging process," he said. "This new experimental method is fast, inexpensive and can be applied to many different targets, from bacterial pathogens to human proteins."

In order to try to boost the efficiency of the crystallization process, Joachimiak and his colleagues at the MCSG and SGC inserted a protease—a certain type of enzyme that breaks down the bonds that connect a protein's amino acids.

Once added, the protease preferentially bound to the proteins at the disordered regions, snipping off the loose ends like a molecular barber. The researchers successfully crystallized and examined nine of these newly shorn proteins that previously had resisted attempts to study them using X-ray crystallography.

"This simple technique offers an opportunity to uncover and characterize the structures of dozens of proteins that up until now we had to study using much more laborious and expensive approaches," Joachimiak said.

This process, known as "limited in situ proteolysis," represents one of several potential "salvage pathways" that biophysicists could use to create more usable protein crystals and reduce waste, Joachimiak said. Currently, scientists' efforts to manufacture and then study a workable crystal on Argonne's Advanced Photon Source yield structural data only about 15 percent of the time. By using proteases to digest part of the protein sample, the Argonne scientists achieved a six percent boost in efficiency.

Joachimiak cautioned that scientists do not have a way to successfully crystallize every protein, even with the use of proteolysis. "There will still be some that are resistant," he admitted, "but we are making enormous strides in our understanding of how exactly these essential substances work."

A research paper, "In situ proteolysis for protein crystallization and structure determination," that detailed the study appeared in the December 4 issue of Nature Methods. The study's X-ray data were collected at the SBC beamlines at the Advanced Photon Source. The MCSG and SGC represent a collaboration of Argonne scientists as well as scientists from Canada and Europe.

Source: Argonne National Laboratory

Explore further: Mapping the protein universe

Related Stories

Mapping the protein universe

October 9, 2015

To understand how life works, figure out the proteins first. DNA is the architect of life, but proteins are the workhorses. After proteins are built using DNA blueprints, they are constantly at work breaking down and building ...

Chemists solve major piece of cellular mystery

August 27, 2015

Not just anything is allowed to enter the nucleus, the heart of eukaryotic cells where, among other things, genetic information is stored. A double membrane, called the nuclear envelope, serves as a wall, protecting the contents ...

Recommended for you

From a very old skeleton, new insights on ancient migrations

October 9, 2015

Three years ago, a group of researchers found a cave in Ethiopia with a secret: it held the 4,500-year-old remains of a man, with his head resting on a rock pillow, his hands folded under his face, and stone flake tools surrounding ...

Mexican site yields new details of sacrifice of Spaniards

October 9, 2015

It was one of the worst defeats in one of history's most dramatic conquests: Only a year after Hernan Cortes landed in Mexico, hundreds of people in a Spanish-led convey were captured, sacrificed and apparently eaten.

Ancient genome from Africa sequenced for the first time

October 8, 2015

The first ancient human genome from Africa to be sequenced has revealed that a wave of migration back into Africa from Western Eurasia around 3,000 years ago was up to twice as significant as previously thought, and affected ...


Please sign in to add a comment. Registration is free, and takes less than a minute. Read more

Click here to reset your password.
Sign in to get notified via email when new comments are made.