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Extra tool in DNA’s repair kit

VANDERBILT (US) — On a good day about one million bases in the DNA in a human cell are damaged. Now researchers have discovered a fundamentally new way that DNA-repair enzymes detect and fix that damage.

The findings are reported in the journal Nature.

Tucked within its double-helix structure, DNA contains the chemical blueprint that guides all the processes that take place within the cell and are essential for life. Therefore, repairing damage and maintaining the integrity of its DNA is one of the cell’s highest priorities.

“There is a general belief that DNA is ‘rock solid’—extremely stable,” says Brandt Eichman, associate professor of biological sciences at Vanderbilt University, who directed the project. “Actually DNA is highly reactive.”

Lesions to bases in DNA are caused by a combination of normal chemical activity within the cell and exposure to radiation and toxins coming from environmental sources including cigarette smoke, grilled foods, and industrial wastes.

“Understanding protein-DNA interactions at the atomic level is important because it provides a clear starting point for designing drugs that enhance or disrupt these interactions in very specific ways,” says Eichman. “So it could lead to improved treatments for a variety of diseases, including cancer.”

Toxic toll
The newly discovered mechanism detects and repairs a common form of DNA damage called alkylation. A number of environmental toxins and chemotherapy drugs are alkylation agents that can attack DNA.

When a DNA base becomes alkylated, it forms a lesion that distorts the shape of the molecule enough to prevent successful replication. If the lesion occurs within a gene, the gene may stop functioning.

To make matters worse, there are dozens of different types of alkylated DNA bases, each of which has a different effect on replication.

One method to repair such damage that all organisms have evolved is called base excision repair. Special enzymes known as DNA glycosylases travel down the DNA molecule scanning for these lesions.

When they encounter one, they break the base pair bond and flip the deformed base out of the DNA double helix. The enzyme contains a specially shaped pocket that holds the deformed base in place while detaching it without damaging the backbone. This leaves a gap (called an “abasic site”) in the DNA that is repaired by another set of enzymes.

Human cells contain a single glycosylase, named AAG, that repairs alkylated bases. It is specialized to detect and delete “ethenoadenine” bases, which have been deformed by combining with highly reactive, oxidized lipids in the body.

However, AAG also handles many other forms of akylation damage. Many bacteria, however, have several types of glycosylases that handle different types of damage.

“It’s hard to figure out how glycosylases recognize different types of alkylation damage from studying AAG since it recognizes so many,” says Eichman. “So we have been studying bacterial glycosylases to get additional insights into the detection and repair process.”

Key discovery
That is how they discovered the bacterial glycosylase AlkD with its unique detection and deletion scheme. All the known glycosylases work in basically the same fashion: They flip out the deformed base and hold it in a special pocket while they excise it.

AlkD, by contrast, forces both the deformed base and the base it is paired with to flip to the outside of the double helix. This appears to work because the enzyme only operates on deformed bases that have picked up an excess positive charge, making these bases very unstable.

If left alone, the deformed base will detach spontaneously. But AlkD speeds up the process by about 100 times. Eichman speculates that the enzyme might also remain at the location and attract additional repair enzymes to the site.

Cancer connection
The new repair mechanism may prove to be the key to understanding the differences in the way that the repair enzymes identify and repair toxic and mutagenic lesions.

That is important because mutagenic lesions that the repair mechanisms miss are copied to daughter cells and so can spread whereas the deleterious effects of toxic lesions are limited to the original cell.

Understanding these differences could lead to more effective chemotherapy agents, Eichman points out. These drugs are strong alkylating agents designed to induce lesions in a cancer patient’s DNA. Because cancer cells are reproducing more rapidly than the body’s normal cells, the agent kills them preferentially.

However, in addition to toxic lesions that kill the cell, the agent also produces lesions that cause mutations, which can lead to additional complications.

Researchers from Penn State and the University of Pittsburgh contributed to the study, which was supported by grants from the American Cancer Society, National Institutes of Health and U.S. Department of Energy.

More research news from Vanderbilt: www.vanderbilt.edu/exploration

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  1. George Birbilis

    > AlkD, by contrast, forces both the deformed base and the base it is paired with to flip to the outside of the double helix. This appears to work because the enzyme only operates on deformed bases that have picked up an excess positive charge, making these bases very unstable.

    this could explain maybe how magnetic fields (say from electrical transmission lines or from electromechanical devices) can cause damage (e.g. leuchemia at young children)

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