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Scientists map three-dimensional protein that creates therapeutic compounds

Press Release Number: 
Friday, June 17, 2005
By Monica Bobra

Using the Stanford Synchrotron Radiation Laboratory (SSRL) at the Department of Energy’s Stanford Linear Accelerator Center (SLAC), three scientists at The Salk Institute for Biological Studies discovered the three-dimensional structure of a protein that bacteria use to make anti-cancer, anti-viral, anti-inflammatory, and anti-oxidant compounds. By effectively engineering this protein, scientists may be able to create new drugs with therapeutic properties. The Salk Institute scientists reported their results in the June 16 issue of the journal Nature.

The bacterial protein, known as Orf2, contains a previously unknown structure shaped like a barrel. Scientists discovered this structure using a process called X-ray crystallography, a technique in which X-rays, produced by synchrotron radiation, hit the protein’s atoms and diffract to produce a two-dimensional pattern. Computers then interpret a set of such patterns and construct a high-resolution three-dimensional image. Researchers gathered data at SSRL, the National Synchrotron Light Source (NSLS) at Brookhaven National Laboratory and the European Synchrotron Radiation Facility.

Schematic diagram of the Orf2 protein

“Looking at the protein’s structure is like opening up a clock and understanding how all the parts fit together and work in unison,” said co-investigator Joseph Noel. “With X-ray instruments from our lab at the Salk Institute, we looked at Orf2 from 30 feet away. With SSRL, we can look at it with a magnifying glass if we need to. That’s an essential part of the whole process.”

Figure 1. A schematic diagram of the Orf2 protein allows scientists to visualize it in three-dimensional space. The teal helical struture is known as a PT barrel. Grey strands of dihydroxynaphthalene (DHN) and geranyl S-thiolodiphosphate (GSPP) reside within the PT barrel; the gold sphere in the protein's core represents magnesium.

Orf 2 is one of a small number in a recently isolated family of proteins that create compounds with anti-microbial, anti-cancer, anti-viral, anti-inflammatory, and anti-oxidant properties. Orf2 accomplishes this remarkable task by adding hydrocarbons known as prenyl groups to flat benzene-like molecules. By manipulating Orf2 using knowledge of its three dimensional shape, scientists could engineer therapeutic drugs based upon naturally occurring chemicals including polyketides, plant flavonoids and stilbenes.

“When we first looked at it, very quickly, we thought: This is one of the most common 3-D folds known in nature,” said Noel, referring to a common protein structure known as a TIM barrel. Soon after, the team realized Orf 2’s structure was incredibly distinctive: its amino acids, which form the protein’s building blocks, are arranged in an entirely different manner.

“We currently use the protein as a surrogate chemist, allowing it to catalyze chemical reactions that would be difficult or impossible to do with traditional chemistry,” said Noel. This allows the researchers to slightly modify existing chemicals, which may lead them to discover new drugs. “We hope to use and engineer the protein to create novel compounds,” said co-investigator Stéphane Richard.

In addition, studying Orf2’s structure “allows us to understand the fundamental process of molecular evolution,” said Noel. The team is analyzing the amino acid sequence of closely related proteins to understand how Orf2 has changed over time. “Using laboratory techniques for genetic manipulation, we can now make changes to Orf2 and its relatives in a manner resembling the path that evolution has taken over the last half a billion years or so,” said Noel.

Without using x-ray crystallography techniques at SSRL, NSLS, and the European Synchrotron Radiation Facility, the scientists couldn’t have identified the more subtle features of Orf2 and noted the key differences between Orf2’s structure and a TIM barrel structure. SSRL and NSLS are scientific user facilities operated by the U.S. Department of Energy Office of Science. NIH provides additional support for the biological crystallography programs at both laboratories.

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[Note: Published in Nature [Nature 435, 983-987 (16 June 2005) | doi: 10.1038/nature03668]; Tomohisa Kuzuyama, Joseph P. Noel and Stéphane B. Richard; "Structural basis for the promiscuous biosynthetic prenylation of aromatic natural products"

Data was collected at the National Synchrotron Light Source (Brookhaven National Laboratory), beamlines X8C and X6A; ESRF (European Synchrotron Radiation Facility), beamline FIP/BM30A; and Stanford Synchrotron Radiation Laboratory (Stanford Linear Accelerator Center), beamline 9.1. Work performed at SSRL was supported by grants from the National Institutes of Health, National Center for Research Resources, Biomedical Technology Program and the Department of Energy, Office of Biological and Environmental Research. Brookhaven National Laboratory and Stanford Synchrotron Radiation Laboratory are funded by the U.S. Department of Energy's Office of Science-]