麻豆淫院

When the cell鈥檚 DNA proofreading system encounters a mismatch in the genome, a number of repair proteins are thrown into action, ripping up the mistake and filling in the correct nucleotides. One of these proteins, an endonuclease called MutL in bacteria, nicks the error-containing strand of DNA with the help of ATP, the energy currency of the cell. However, the exact mechanism by which ATP regulates the DNA-cutting enzyme has eluded biochemists.

Now, a research team from Japan has shown that ATP physically binds the repair protein, causing a structural rearrangement at the enzyme鈥檚 catalytic site that triggers the genetic slicing and dicing.

Led by Kenji Fukui from the RIKEN SPring-8 Center in Harima, the researchers performed a series of structural imaging experiments in the heat-loving bacterium Aquifex aeolicus to tease apart how ATP affects MutL (Fig. 1). Using a mass spectrometry technique designed for monitoring conformational changes in proteins, they showed that ATP first binds to one end of the MutL protein. Through a series of physical interactions, this binding then induces structural changes at the opposite end of the protein鈥攚here the catalytic site is located鈥攕purring the enzyme into action.

Importantly, the study revealed two novel, highly conserved regions of MutL鈥攐ne in the ATP-binding region, another in the catalytic region鈥攖hat are essential for the protein鈥檚 DNA cleaving activity. 鈥淲e were surprised because the newly identified catalytic region had not been thought to be involved in the catalysis of endonucleolytic reaction,鈥� says Fukui.

In the human equivalent of MutL, mutations in this catalytic region are known to cause Lynch syndrome, a heritable form of cancer marked by a faulty DNA mismatch repair system. Consistent with this observation, Fukui and colleagues studied MutL with mutations at the catalytic region and saw a decrease in enzymatic activity. As such, the team鈥檚 study could provide a molecular explanation for the repair defect seen in people with that disease. 鈥淒ysfunction of MutL endonuclease activity [seems to] be one of the major causes for Lynch syndrome,鈥� Fukui notes.

More fine-scale structures are still needed, however, to fully characterize the ATP-dependent structural rearrangement of the protein in both healthy and mutated forms of MutL. To that end, Fukui is now further probing the protein using x-ray crystallography and other methods. 鈥淭he precise biochemical characterization of A. aeolicus鈥檚 MutL will accelerate the cellular biological studies on human MutL homolog,鈥� Fukui says.