The team's study reveals details of the new DNA repair pathway

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The team's study reveals details of the new DNA repair pathway

A team of Vanderbilt researchers discovered how a DNA repair pathway protein protects sites from damage to avoid mutations and maintain genome integrity.

"DNA repair is essential to prevent cancer. The loss or ineffectiveness of DNA repair leads to mutations and changes in our chromosomes that lead to the development of cancer," said Dr. David Cortez, Professor of Cancer Research and Professor of Biochemistry at Ingram.

Earlier this year, Cortez and his colleagues reported a new DNA repair mechanism involving a protein called HMCES. Now, in collaboration with Dr. Brandt Eichman and his group, researchers have discovered structural details on how HMCS binds to damaged DNA to protect it from mutation. Their findings were published in the journal Nature Structural & Molecular Biology.

"This is not only a great scientific story, but also a great collaboration between two graduate students," said Eichman, William R. Kenan, Jr. Professor and Head of the Department of Biological Sciences.

The Cortez team has already shown that HMCS has a role to play in repairing "outliers" - the most common type of damaged DNA - in single-stranded DNA. Abominable sites occur up to 20,000 times a day in human cells. When HMCS is missing, cells accumulate DNA damage and have increased genetic stability.

In studies led by postdoctoral fellow Kareem Mohni, the researchers demonstrated that HMCS binds to an abasal site and forms a DNA-protein cross-linkage.

Petria Thompson, a graduate student at Cortez Laboratory, wanted to better understand the chemical nature of the bond between HMCS and an astonishing site. Using biochemical experiments, she showed that the crosslinking was extremely stable and developed methods to purify large amounts of the DNA-protein crosslinking.

Thompson then teamed up with Katherine Amidon at the Eichman Laboratory for structural studies.

The pair purified DNA-protein crosslinks using two proteins: human HMCES and an E. coli homologous protein. Starch produced high quality crystals of both DNA-bound proteins and used X-ray crystallography to determine a high resolution structure of the DNA-protein cross-linking.

The type of chemical bonds they have identified between the protein and the abase site explains the remarkable stability of DNA-protein crosslinking.

Thompson and Starch "worked on all aspects of protein purification, crystallization and biochemical experiments by sharing their expertise, data and reagents," said Dr. Eichman. "This collaboration has allowed the project to progress much more effectively than it would have otherwise."

The structure revealed that HMCS has specificity for abase sites in single-stranded DNA at junctions that occur during replication - when a protein polymerase that copies DNA meets an abase site. The results support HMCS's role in protecting abasal sites during DNA replication.

"Based on our previous biochemical studies, we predicted that HMCS would have a binding preference for these types of abominable sites," said Cortez. "It is exciting that structural biology studies have confirmed our predictions.

How the repair of the abasal site occurs after the formation of the DNA-protein cross-linkage is an open question and will be the subject of future studies.