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Bacterial Protein Structure Hints at Mechanism of a Class of Premature Aging Diseases

Bacterial Protein Structure Hints at Mechanism of a Class of Premature Aging Diseases
Bacterial Protein Structure Hints at Mechanism of a Class of Premature Aging Diseases


Duke Health News Duke Health News

DURHAM, N.C -- Using a bacterial model system, scientists at Duke University Medical Center have determined the first step in understanding how a new class of genetic flaws may translate into diseases.

Deepak Bastia and Steven White, professors of microbiology at Duke, have determined how a protein, called a replication termination protein (RTP), stops DNA replication in the bacterium Bacillus subtilis, a common experimental organism. Rather than simply acting as a roadblock to replication, as had been previously thought, RTP is more like a one way door, allowing the DNA replication machinery to move in only one direction along the DNA strand, the researchers said. They said the work has implications for human disease because RTP interacts with a class of proteins known as helicases, which have been shown to be involved in several genetic diseases. Bastia is now starting work on identifying the comparable protein in humans. The researchers' findings appear in the Nov. 29 issue of the journal Cell. The research was funded by the National Institutes of Health.

RTP has been known to exist for 20 years, but it wasn't until two years ago that its structure was revealed by Bastia and White. "This [recent] work relates structure to function and takes us one step further in understanding how RTP works," Bastia said.

When DNA replicates, it unzips, starting and ending at specific sites. The helicases are responsible for pulling apart the DNA strands, which are twisted in a helical pattern. "RTP ensures that when DNA replicates, it is brought to a neat and accurate finish, and the cell knows that it is time to divide into two," White said.

Recently, helicase malfunction has been found responsible for a class of genetic diseases including Werner's syndrome and Cockayne's syndrome, premature aging syndromes; Bloom's syndrome, a type of dwarfism; and some kinds of skin cancer. Understanding of these diseases on the molecular level is minimal. But since the helicase's job is to help replicate the chromosomes, any malfunction would be expected to have severe consequences. "No one knows anything about arresting helicases in the human system," Bastia said. "There's zero information in the literature."

The events taking place between the genetic flaw and its expression as a disease are unknown. "It's not enough to know where a defect in the gene is." Bastia said. "You want to know how it causes the disease."

If the mechanism of a genetic malfunction can be mapped out, said Bastia, drug therapies will be easier to design as would gene therapies. The new research is the first step in that direction. The Duke researchers have shown that the RTP, rather than simply acting as a road block to replication, actually interacts with the helicase protein.

"That's a major controversy in this field," White said. A competing theory, published in the Oct. 17, 1996, issue of the journal Nature is that RTP mechanically interferes with helicases in a non- specific way, but the Duke research shows that RTP actually interacts with the helicase on a molecular level. "There's a flexible region on the helicase that the RTP seems to interact with," said White.

To test how RTP works, the researchers had to create a mutant version of the protein that would not interact with the helicase but wouldn't be deformed beyond recognition.

"Our big worry was that we would have destroyed the structure," said White, who determined RTP's structure two years ago using X-ray crystallography. Engineering mutations at the molecular level, the scientists changed one amino acid in RTP to test their hypothesis that it interacts with a specific region on the helicase.

"Identifying the corresponding proteins from a human system and then finding a mutant protein will lead to understanding of the disease process involving helicase proteins," Bastia said. "And this understanding may eventually help to work out gene therapy techniques."

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