Tool belt protein links molecular ‘motor’ and ‘typewriter’ to keep DNA replication on track

12 June 2014

Crystal structure of the ‘toolbelt’ protein seen from the top, with the three docking sites visible.

Image: Crystal structure of the ‘toolbelt’ protein seen from the top, with the three docking sites visible.

Scientists at Cancer Research UK's London Research Institute (LRI; now part of the Francis Crick Institute) have described the structure of a protein that plays a key role in the DNA replication process.

The protein physically links a motor that unwinds the DNA double helix and a molecular 'typewriter' that sythesises a new DNA strand.

The work has implications for understanding and intervening in cancer, which can happen when the motor and typewriter stop working together properly.

Dr Alessandro Costa of LRI explained: "Each proliferating cell in our body needs to generate an accurate copy of its own chromosomes, before dividing. This task is performed by the replisome, a complex molecular machine.

"Within this replisome, the activities of the motor and typewriter need to be accurately orchestrated to preserve the physical integrity of our genome. Despite the crucial importance of the DNA replication process, we still have very little knowledge on how the replisome components interact with one another in our cells to copy DNA."

The team isolated individual replisome proteins and mixed them together to reconstitute a macromolecular assembly. They then used an electron microscope to image tens of thousands of individual molecules and computational methods to statistically analyse and average these molecules. Combining this information with protein crystallography allowed them to generate a structural model to explain how DNA unwinding and synthesis is coordinated in cells. Finally, the model was confirmed in a living organism.

They found that the linking protein works like a tool belt, made of three identical units that come together to form a ring, providing three docking sites. Both the DNA unwinding motor and the molecular typewriter possess a short 'tail' with an identical sequence of amino acids - these work like hooks that latch onto the tool belt.

Dr Costa said: "Uncoupling the activities of the DNA unwinding motor and the DNA typewriter can have severe consequences. For example, if the typewriter stops working while the motor keeps unwinding DNA this causes the accumulation large stretches of single-stranded DNA, which is much more fragile than the DNA double helix. This can lead to genomic instability, which is a hallmark of cancer cells.

"By learning how the DNA unwinding and synthesis machineries work together during genome replication, we can find the tools to intervene when things go wrong."

Next the team plans to investigate what the third space on the tool belt is for - and whether this third protein acts when the replisome encounters damaged DNA.

The paper, A Ctf4 trimer couples the CMG helicase to DNA polymerase α in the eukaryotic replisome, is published in Nature.

  • New research has uncovered the structure of a protein that physically links amotor that unwinds the DNA double helix and the molecular 'typewriter' that sythesises a new DNA strand in the DNA replication process.
  • Correct DNA replication by cells is important because cancer can occur if the process goes awry.
  • The work was carried out by scientists at Cancer Research UK's London Research Institute, the University of Cambridge and the Medical Research Council Protein Phosphorylation and Ubiquitylation Unit at the University of Dundee.