Baker and Heather Rock Woods
In the world of molecules, DNA tends to get top billing at the expense
of RNA, which is critical for turning DNA’s genetic blueprint into working
proteins. Researchers at the Stanford University School of Medicine have
published significant insights into how the RNA molecule completes this
task in two back-to-back papers in the February 13 issue of Science
A schematic of SSRL’s robotic screening system which
finds the best crystals to study.
(Image Courtesy of Roger Kornberg)
All the genetic information contained in DNA is silent, said Roger
Kornberg, Stanford professor of medicine and structural biology. What
gives it a voice is RNA polymerase, the enzyme that copies DNA into RNA
through a process called transcription. Along with more than a dozen
helper molecules, RNA polymerase determines which proteins are produced
within a cell. But before scientists can understand the transcription
process, they must first unveil the inner structure of RNA polymerase,
which is where SSRL comes in.
Kornberg’s lab has been studying RNA and the enzyme that makes it for
more than 20 years. Past studies from the lab have shown that the
machinery of the RNA polymerase system is in three layers. His group
published groundbreaking findings in 2001 outlining the structure of the
innermost layer. The recently published papers focus on the middle layer,
which contains many of the helper molecules.
To see the structure of the protein layers, the group passed SSRL’s
extremely bright x-rays through a crystallized version of the proteins.
The crystal scatters the x-rays, generating a distinctive diffraction
pattern that reveals the sample’s three-dimensional atomic structure in
To find good diffracting crystals out of the hundreds made, the
researchers used a new automatic robotic screening system developed at
SSRL with grants from the National Institutes of Health. The automated
screening system stores the tiny frozen crystals on nylon loops at the end
of metal pins. A robotic arm retrieves each pin and aligns the crystal in
the path of the X-ray beam. The robot can automatically test 300 samples
without the need for researchers to manually transfer each sample as was
done in the past. The new robots are becoming operational on all of SSRL’s
crystallography beam lines.
"It saves a lot of time while optimizing the quality of the data," said
SSRL scientist Mike Soltis, head of the macromolecular crystallography
group. "With the new system, the Kornberg group screened 130 crystals in
seven hours without losing any. Two weeks earlier, they had manually
mounted 100 crystals in 24 hours, losing a few crystals and much sleep in
At the level of detail the researchers obtained, some intriguing
structures came to light, offering the first real understanding of the
defining events of transcription. They saw a docking site that might
reveal the starting point of transcription, a spot where the RNA
polymerase is correctly situated on a gene. They also saw something
completely unexpected: a "finger" of the helper molecule that pokes into
the polymerase’s active center. The researchers speculate that the poking
action may help slow down the transcription process so that the strands of
DNA and newly made RNA can separate properly.
"This turned out to be quite interesting. No one had even speculated
about it before," said David Bushnell, a research associate and first
author of one of the papers. "We think the protrusion reaching into the
enzyme makes sense of a lot of genetic and biochemical data that people
were scratching their heads over."
Catching the Polymerase in Action
The second paper describes how the team caught a snapshot of the
polymerase in action, something that hadn’t been done before. Kenneth
Westover, an MD/PhD student and first author of the second paper,
developed a method in which the newly made RNA could be visualized
separating from the DNA.
How the strands of RNA and DNA are pushed apart has a simple physical
explanation: the RNA polymerase inserts itself as a wedge between the two,
with the RNA trailing out an opening in the polymerase. That same opening
is the one that the protein finger dips into.
"These two papers are both quite astonishing in what they reveal,"
Mitzi Baker is a science writer at Stanford’s School of Medicine.