Cryo-EM reveals ignition mechanism for DNA replication

IMAGE: This is Mcm2-7 double hexamer on DNA, a critical early step in DNA replication.

Image credit: 
Courtesy of Van Andel Research Institute and Dr. Huilin Li.

GRAND RAPIDS, Mich. (Oct. 23, 2017) -- An international team, led by scientists at Van Andel Research Institute, Cold Spring Harbor Laboratory and Imperial College London, has shed new light on a critical step in DNA replication, offering fresh insights into a fundamental process of life and driver of many different diseases, including cancer.

"Previous studies have described how enzymes assemble and gather around DNA to prepare it for replication. Here, we describe what these enzymes do to DNA once they are in place," says Huilin Li, Ph.D., a professor in the Center for Epigenetics and one of the senior authors on a study published today in Proceedings of the National Academy of Sciences.

The work is part of a long-time collaboration between Li, Bruce Stillman, Ph.D., president of Cold Spring Harbor Laboratory, and Christian Speck, Ph.D., professor at Imperial College London.

DNA replication is a tightly choreographed process that copies the genetic code, allowing its instructions to be passed on from one generation of cells to the next. In diseases like cancer, these mechanisms can fail, leading to uncontrolled or faulty replication with devastating consequences.

"How this complex process starts is not well understood at the molecular level," Li adds. "Our hope is that the more mechanistic detail we learn about how replication works, the better able others will be in developing new treatments for cancer and other diseases."

Their discovery wouldn't have been possible without a revolutionary technology called cryo-electron microscopy, or cryo-EM, which allows scientists to see critical biological components in exquisite, atomic detail. Using the powerful microscopes in VARI's David Van Andel Advanced Cryo-Electron Microscopy Suite, Li and his collaborators revealed the very first steps in DNA replication--the separation of the two DNA strands from one another.

Before replication can take place, a pair of structures called heterohexameric minichromosome maintenance (Mcm2-7) helicases are assembled head to head on the DNA double helix as a double hexamer. They eventually separate into two functional helicases and, in the process, each push out one strand of the double helix. Later, when DNA replication starts, the two helicases each move on one strand of DNA in opposite directions to unwind the helix.

Computational rendering of the cryo-EM images revealed the three-dimensional structure of these helicase enzymes. Imaging from the team's study shows the helicase enzymes binding to 60 base pairs of the DNA double helix.

Li likens it to a spring-loaded mechanism that puts pressure on either side of DNA, bending the helix into a zig-zag shape. This positions the DNA strands toward two side-way gates, ready to be pushed out in the next stage when the two Mcm2-7 hexamers disjoin, going opposite directions to "unzip" the double helix.

"These are processes at the very foundation of life that have largely remained a mystery to biologists since the discovery of DNA double helix more than 60 years ago," says Li. "Thanks to technologies like cryo-EM, we are able to 'see' the operational mechanism in action, which gives us valuable knowledge to improve health for people around the world."

Van Andel Research Institute