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: Closer look at microorganism provides insight on carbon cycling

Related Stories

Self-assembly of molecular Archimedean polyhedra

July 1, 2015

Chemists truly went back to the drawing board to develop new X-shaped organic building blocks that can be linked together by metal ions to form an Archimedean cuboctahedron. In the journal Angewandte Chemie, the scientists ...

Disabling antibiotic-resistant bacteria

June 18, 2015

Dreaded bacterial-related diseases have killed untold numbers of humans over the centuries. Today, two million illnesses and nearly 23,000 deaths can be attributed to antibiotic-resistant bacteria throughout the United States, ...

3D potential through laser annihilation

June 16, 2015

Whether in the pages of H.G. Wells, the serial adventures of Flash Gordon, or that epic science fiction saga that is Star Wars, the appearance of laser beams—or rays or phasers or blasters—ultimately meant the imminent ...

The most complete functional map of an entire enzyme family

May 25, 2015

Researchers at two Department of Energy-funded Scientific User Facilities collaborated with one of three Bioenergy Research Centers to develop and analyze high-resolution crystal structures of an enzyme from the cellulose-degrading ...

New Argonne study may shed light on protein-drug interactions

January 15, 2008

Proteins, the biological molecules involved in virtually every action of every organism, may themselves move in surprising ways, according to a recent study from the U.S. Department of Energy’s Argonne National Laboratory ...

Recommended for you

French teen finds 560,000 year-old tooth (Update)

July 28, 2015

A 16-year-old French volunteer archaeologist has found an adult tooth dating back around 560,000 years in southwestern France, in what researchers hailed as a "major discovery" Tuesday.

0 comments

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.