Research team models moving ‘pucks’ that help DNA replicate

Patterns of rice moving

A helicase protein model created at Rice University shows a before and after of how the six-sided ring moves along DNA to split double-strands into single-strands in response to hydrolysis of DNA. ‘ATP during replication. 1 credit

Knowing the structure of a complex biological system is not enough to understand its functioning. It helps to know how the system is moving.

With this in mind, researchers at Rice University have modeled a key mechanism by which DNA replicates.

Combining structural experiments and computer simulations, bioscientist Yang Gao, theoretical physicist Peter Wolynes, graduate student Shikai Jin and their colleagues discovered details of how helicases, a family of ring-shaped motor proteins, disrupt DNA during replication. Their work could reveal new targets for drugs that fight disease.

The synergy between experiments and large-scale simulations that they describe in the Proceedings of the National Academy of Sciences could become a paradigm for modeling the mechanisms of many complex biological systems.

“These are dynamic processes that cannot be captured well with experimental methods alone,” said Gao, assistant professor of biosciences and CPRIT researcher in cancer research. “But it is important to show the mechanisms of these helicases, because they are essential for DNA replication, and also possible targets for drugs.”

Hexameric helicases have six sides that self-assemble from peptides into a washer-shaped ring that separates parental double strands of DNA into daughter single strands. So far, researchers have not been able to determine how the helicase progresses as it unzips the double strand.

Rice’s simulations support the idea that the DNA-binding loops in the six helicase subunits form a sort of staircase that descends the DNA backbone fueled by ATP hydrolysis. , the process by which the chemical energy stored in ATP molecules is released.

It was known that ATP is attracted to the NTPase proteins in each subunit to drive stepping motion. But the researchers hadn’t precisely understood that the hydrolysis of ATP is the key. The Rice team found that ATP binds helicase subunits tightly, but hydrolysis dramatically lowers the energy barrier for subunit dissociation, allowing the protein to move forward.

A simulation shows how a six-sided helicase protein moves along a DNA strand as it separates double strands into single strands during replication. Rice University theorists have discovered that the hydrolysis of ATP is the key to the staircase movement of proteins. A complete step can be seen here. Credit: Yang Gao and Shikai Jin

The researchers noted that because the helicase-DNA complex is so large, there have only been a few attempts to simulate the translocation of helicase from one end of the strand to the other. Rice’s hybrid of two coarse-grained simulation techniques allowed the process to be studied from start to finish.

The simulations revealed several previously unknown intermediate states and highlighted the interactions involved in the long-range movement of helicase. They showed that each translocation step can travel more than 12 nucleotides along the backbone.

To search for the mechanism, the team focused on bacteriophage T7, a virus that infects bacteria often used as a model system. To simulate his helicase, known as gp4, the researchers combined two force fields: AWSEM, originally developed by Wolynes and colleagues to predict protein folding, and open3SPN2, a DNA simulator developed by the engineer molecular scientist Juan de Pablo of the University of Chicago. Force fields describe the forces that dictate how atoms and molecules move when in contact. (The new forcefield combination was the subject of a 2021 article edited by Rice in Computational Biology PLOS.)

Both force fields are coarse-grained molecular models based on machine learning that use only a subset of atoms in a system, but still provide accurate results while dramatically reducing computation time.

“Combining the models enabled GPU acceleration so we could make our molecular dynamics simulations very fast,” Jin said. “The combined software is now 30 times faster than the versions we’ve used for other studies.”

It helps that T7 is half the size of helicases in human cells. “In human systems, there are six different polypeptide chains in the helicase, but in T7 it’s the same one that makes six copies,” he said.

“Because our new form of open3SPN2 deals with a single strand of DNA, it allows us to analyze processes where usually double-stranded DNA opens as it does in the presence of helicase,” said said Wolynes, co-director of Rice’s Center for Theoretical Biological Physics. “The single-stranded DNA force field itself is new, but it was only a background in this project, where it allows us to look at the process in detail.”

“The cryo-EM structures we have for these essential complexes are physiologically precise, but these systems are dynamic,” Gao said. “They have to move to do their job, and there’s still a lot we want to know about how they do it.

“That’s where these computational models can make a big contribution and they will, for sure, be adapted to other large systems to look at rather important questions,” he said.

Researchers identify how bacterial replicative helicase opens to start DNA replication process

More information:
Shikai Jin et al, Computational exploration of the mechanism of bacteriophage T7 gp4 helicase translocation along ssDNA, Proceedings of the National Academy of Sciences (2022). DOI: 10.1073/pnas.2202239119

Provided by Rice University

Quote: Research team models moving “washers” that help DNA replicate (2022, August 9) Retrieved August 10, 2022 from dna-replicate.html

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Research team models moving ‘pucks’ that help DNA replicate

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