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How an Enzyme Repairs DNA via a “Pinch-Push-Pull” Mechanism

SDSC’s Comet Provides Computing Power to Model Mechanism Critical to Maintain Stability of DNA

Published May 30, 2018

Thymine DNA glycosylase (TDG) in complex with DNA is shown on the top left panel. Free energy profile for the TDG-induced base flipping is shown in the top right panel. Structures of the flipped out DNA base interacting the enzyme at various intermediate states are shown and mapped onto the base flipping path (as indicated by arrows).  Credit: Dr. Chunli Yan and Tom Dodd, Georgia State University

At first, the alliterative phrase sounds remarkably simplistic: “pinch-push-pull.”

But when it comes to preserving one of biology’s basic processes – the replication of the genetic code – this single-syllabic triplet spells the difference between normal cell division and a potentially hazardous mutation.

In a study published in the May 21, 2018 issue of the Proceedings of the National Academy of Sciences, a team of researchers – aided with supercomputing resources from the San Diego Supercomputer Center (SDSC) based at UC San Diego – created a dynamic computer simulation to delineate a key biological process that allows the body to repair damaged DNA.

The simulation describes in detail how a key DNA repair enzyme called thymine DNA glycosylase (TDG) first searches for damaged DNA among a vast background of normal DNA. Once discovered, the enzyme compresses or pinches the DNA so the lesion, a chemically modified or mismatched DNA base, flips out of the DNA base stack and is then pushed into the enzyme’s active site.  Once the aberrant base is captured, the enzyme pulls and snips it away from the rest of the DNA molecule. This initiates subsequent biochemical steps to restore DNA to its original condition, thus completing the repair.

Or, as the paper summarizes: “pinch-push-pull.”

“Our paper describes a novel mechanism for how this lesion search, base interrogation, and flipping occur and explains the origin of the extraordinary specificity of thymine DNA glycolylase,” said  Ivaylo Ivanov, associate professor of chemistry at Georgia State University and the study’s principal investigator. “Deeper understanding of these processes and the respective biochemical pathways could be important medically as well as biologically.”

In essence, TDG has a dual role in the DNA repair process: to maintain the integrity of DNA, and remove epigenetic markers – primarily oxidized derivatives of 5-methyl cytosine – that can silence a gene’s activity without changing its sequence. Put another way, TDG works as a molecular scissor that excises a damaged DNA base or a base carrying an epigenetic modification.

“Understanding the selectivity of TDG is important,” added Ivanov. “These repair proteins are targets for inhibitor development and these can be used as adjuvants in cancer therapy.”

Much of this study builds on ground-breaking discoveries in DNA repair which garnered the 2015 Nobel Prize in Chemistry for Tomas Lindahl, Paul Modrich, and Aziz Sancar for “having mapped and explained how the cell repairs its DNA to safeguard genetic information.”

From a chemical perspective, DNA is a remarkably stable molecule.  But damage to DNA in living cells occurs thousands of times per day, largely stemming from two sources: errors in genome replication or environmental agents such as ultraviolet light, toxic chemicals, ionizing radiation, and the internal presence of reactive molecules.

“Considering how often DNA comes under attack each day, it’s amazing how any lifeform manages to preserve and sustain its genetic code,” said Ivanov.

The genome remains relatively intact over years thanks to a host of repair mechanics circulating in our cells. One of the first and most important processes employed by these repair teams is called base excision repair or BER, identified in 1996 by Lindahl, then director of the Clare Hall Laboratory at Cambridge University. In essence, Lindahl discovered that glycosylases represented the first step in the DNA repair process.

Subsequent research further explained in more detail how the process worked, including computer models based on the X-ray analysis of protein crystals that captured snapshots of the mechanism. However, these static images still failed to describe critical intermediary steps.

In their PNAS article, the Georgia State researchers turned to computational molecular dynamics techniques and SDSC’s Comet supercomputer to simulate the gyrations and seemingly spasmodic movements of TDG as it sculpts or compresses its targeted DNA, flips, and excises detected lesions. Allocations for Comet were provided by the eXtreme Science and Engineering Discovery Environment (XSEDE), funded by the National Science Foundation (NSF).

Among other things, the researchers were able to calculate favorable “free energy profiles” guiding TDG in its lesion search.

“Think of a free energy profile as a sort of map,” said Ivanov. “Valleys on the map are favorable places for the extruded base to reside while it transitions toward the active site of the enzyme. The measure of ‘favorable’ in this case is a thermodynamic quantity we call free energy. The path will follow the lowest free energy regions.”

As summarized by the study: “Our results show that DNA sculpting, dynamic glycosylase interactions, and stabilizing contacts collectively provide a powerful mechanism for the detection and discrimination of modified bases and epigenetic marks in DNA.”

Ivanov said their study would have been difficult to complete if the research team had to rely on local computer resources.

Comet was the best XSEDE resource for us because it provided access to GPUs (graphics processing units) and the code that we used (Amber 16) provides one of the fastest GPU implementations of molecular dynamics” he said.

Also participating in this study, titled “Uncovering universal rules governing selectivity of the archetypal DNA glycosylase TDG”, were Thomas Dodd, Chunli Yan, Bradley Kossman, and Kurt Martin, all from Georgia State University.

Support for the research was provided by a grant from the National Institutes of Health (GM110387) and the NSF (MCB-119521). In addition to allocations from XSEDE, computation resources were provided by the National Energy Research Scientific Computing Center (NERSC), supported by the Department of Energy Office of Science.

About SDSC

As an Organized Research Unit of UC San Diego, SDSC is considered a leader in data-intensive computing and cyberinfrastructure, providing resources, services, and expertise to the national research community, including industry and academia. Cyberinfrastructure refers to an accessible, integrated network of computer-based resources and expertise, focused on accelerating scientific inquiry and discovery. SDSC supports hundreds of multidisciplinary programs spanning a wide variety of domains, from earth sciences and biology to astrophysics, bioinformatics, and health IT. SDSC’s petascale Comet supercomputer is a key resource within the National Science Foundation’s XSEDE (Extreme Science and Engineering Discovery Environment) program.