Scientists at SSRL have found an Achilles heel in the brutal diseases anthrax, botulism, syphilis, diarrhea and Lyme’s disease.

These single-celled disease organisms need a protein called thymidylate synthase complementing protein (TSCP) to replicate. TSCP is an enzyme, a type of protein that catalyzes chemical changes in the molecules that bind to it without changing itself. SSRL’s concentrated x-rays revealed the 3-D structure and function of this protein, which has enabled researchers to create a computer model of a molecule that could block it and thus the organisms that rely on it to survive. The research was a cooperative effort with the Joint Center for Structural Genomics.

The molecules dUMP and FAD attach to the binding site of the TSCP enzyme (left), where dUMP is chemically converted into dTMP (a part of DNA necessary for cell replication). If an inhibitor blocks the binding site (right), dUMP and FAD cannot attach, dTMP cannot be made and the cell cannot in theory reproduce. (Image by Nicole Rager)

"The unique structure of the TSCP enzyme and its discovery last year in numerous pathogenic organisms provide an exciting opportunity for drug design," said Irimpan Mathews, a protein crystallographer in the Structural and Molecular Biology Group at SSRL. The protein x-ray crystallography data was published in the June issue of Structure.

Mathews and his colleagues used the unique x-ray pictures of TSCP to determine the key binding sites where small molecules attach to the TSCP enzyme to undergo chemical reactions. TSCP converts a molecule called dUMP into dTMP, an essential part of DNA (dTMP provides the T in the 4-letter DNA alphabet). In order for a cell to divide into two, it makes a copy of its DNA—the cell’s instruction manual—so that each new cell will have one and will know what to do. If the cell lacks dTMP, it can’t build a DNA copy.

The researchers worked with a TSCP enzyme from a bacterium that lives at high temperatures. Their next step was to learn how similar that enzyme is to TSCP enzymes in different organisms.

By comparing the genetic makeup of 61 TSCP enzymes, they found that all known members of this enzyme family share the same structure at their core. This commonality allows one drug to potentially inhibit the activity of all TSCPs – and therefore stop the replication of many disease-causing organisms.

The enzyme is an especially attractive target for therapeutic drugs because humans don’t have TSCP (we use a completely different enzyme to make dTMP). This reduces the chances of a drug interfering with human cells.

Exploiting the enzyme’s unique binding site, Mathews and his colleagues have proposed a design for an inhibitor that would hinder only TSCP. Molecules dock at the binding site like boats in a slip before they can be catalyzed. Conversely, an inhibitor can dock at the binding site first to thwart the chemical reaction.

The TSCP binding site has anchorages for the molecules dUMP, FAD, and a third small molecule. Although each molecule has a different shape, it fits snugly into its attachment area, the way different computer cables – modem, power, USB – have a corresponding port to plug into. The inhibitor is shaped to plug up the attachment areas of both dUMP and FAD, preventing the two molecules from fastening in computer simulations.