These cartoons
show the old "textbook" view of the replisome, left, and the new
view, right,
revealed by
electron micrograph images in the current study. Prior to this study,
scientists
believed the two
polymerases (green) were located at the bottom (or back end) of the helicase
(tan), adding
complementary DNA strands to the split DNA to produce copies side by side.
The new images
reveal that one polymerase is located at the front end of the helicase. The
scientists are
conducting additional studies to explore the biological significance of this
unexpected
location.
(November 2, 2015) State-of-the-art
electron microscope images reveal that structure of DNA-copying protein complex
differs from long-held textbook view
The first-ever images of the protein complex that unwinds,
splits, and copies double-stranded DNA reveal something rather different from
the standard textbook view. The electron microscope images, created by scientists
at the U.S. Department of Energy's Brookhaven National Laboratory with partners
from Stony Brook University and Rockefeller University, offer new insight into
how this molecular machinery functions, including new possibilities about its
role in DNA "quality control" and cell differentiation. The images
and implications are described in a paper published online by the journal
Nature Structural & Molecular Biology, November 2, 2015.
"This work is a continuation of our long-standing
research using electron microscopy to understand the mechanism of DNA
replication, an essential function for every living cell," said Huilin Li,
a biologist with a joint appointment at Brookhaven Lab and Stony Brook
University. "These new images show the fully assembled and fully activated
'helicase' protein complex—which encircles and separates the two strands of the
DNA double helix as it passes through a central pore in the structure—and how
the helicase coordinates with the two 'polymerase' enzymes that duplicate each
strand to copy the genome."
Three-dimensional
structure of the active DNA helicase bound to the front-end
DNA polymerase
(Pol epsilon). The DNA polymerase epsilon (green) sits
on top rather than
the bottom of the helicase.
Studying this molecular machinery, known collectively as a
"replisome," and the details of its DNA-copying process can help
scientists understand what happens when DNA is miscopied—a major source of
mutation that can lead to cancer—or learn more about how a single cell can
eventually develop into the many cell types that make up a multicellular
organism. But no one has produced a real structure of a replisome at any
resolution for any organism until now.
Collaborating
scientists and study coauthors Zuanning Yuan, a graduate student
at Stony Brook
University (standing), Huilin Li of Stony Brook and Brookhaven Lab
(seated, back),
and Jingchuan Sun of Brookhaven Lab (seated, front)
examining protein
structures.
"All the textbook drawings and descriptions of how a
replisome should look and work are based on biochemical and genetic
studies," Li said, likening the situation to the famous parable of the
three blind men trying to describe an elephant, each looking at only one part.
Those textbook drawings show the helicase moving along the DNA, separating the
two strands of the double helix, with two polymerases located at the back where
the DNA strand is split. In this configuration, the polymerases would add
nucleotides (molecules containing the complementary A, T, G, and C bases of the
genetic code) to the side-by-side split ends as they move out of the helicase
to form two new complete double helix DNA strands.