Painting the picture with neutrons

Neutrons see light elements, such as hydrogen, and X-rays see heavier elements, such as carbon, nitrogen and oxygen. Neutron diffraction at SNS and HFIR, in-house X-ray diffraction at ORNL and synchrotron X-ray diffraction at Argonne National Laboratory’s Advanced Photon Source provided insights the team needed to definitively characterize the enzyme’s chemical reaction.

“Neutrons allow us to see hydrogen atoms, and hydrogen drives chemistry,” Drago said. “Enzymes are about 50% made up of hydrogen atoms. In terms of electrostatics, hydrogen also carries a positive charge, which dictates the environment of the enzyme. Once you have a crystal that will diffract neutrons, you have everything you need. You see the positions where hydrogens are located and, equally as important, the positions lacking hydrogens. You get the whole picture.”

As shown in the animation, cancer cell mitochondria overproduce the SHMT enzyme, a tetramer constructed from four identical peptide chains, or protomers, shown in gray. SHMT functions by using pyridoxal-5′-phosphate, covalently bound to SHMT, and tetrahydrofolate, shown in gold and purple, respectively. Tetrahydrofolate acts as a substrate that binds to the active sites of all four protomers. The hydrogen atoms, shown flashing in green, revealed the exact catalytic mechanism and the roles of various amino acid residues in the enzyme’s active sites. Once the enzyme releases tetrahydrofolate, an inhibitor, shown in blue, could be designed to block further chemical reactions at these sites, arresting the one-carbon metabolic pathway in cancer cells.

“The locations of the hydrogen atoms determine protonation states of specific chemical groups inside the enzyme’s active sites,” Kovalevsky said. “Thus, they provide information on the electric charge distribution, or electrostatics. This knowledge is crucial to designing small-molecule inhibitors that would bind to SHMT, replacing tetrahydrofolate and halting the enzyme function.”

This Oak Ridge National Laboratory news article "Neutrons settle 40-year debate on enzyme for cancer drug design" was originally found on https://www.ornl.gov/news