A joint research group led by Ryo Tanifuji and Hiroki Oguri from the University of Tokyo and Yoshikazu Tanaka from Tohoku University has successfully visualized SacC, an enzyme that plays a key role in building the complex five-ring core of safracin B. This core structure is shared with trabectedin, a drug used to treat soft tissue sarcoma. While getting a clear picture of the enzyme in its working state is a remarkable achievement in and of itself, the method pioneered during the project might have the widest reach. With its potential applicability to a broad range of related enzymes that produce medically useful compounds, the approach could open new avenues for research into not only optimizing already existing compounds but also creating new ones. The findings were published in the Journal of the American Chemical Society (JACS).
The production line for the drug trabectedin starts with the bacterial compound safracin B, which the enzyme SacC assembles within the bacteria. The enzyme carries out a remarkable, seven-step catalytic cascade, stitching together two new carbon-to-carbon bonds and producing a five-ring core scaffold in a single pass.
“Synthesizing such a complex scaffold artificially,” explains Tanifuji, “takes many carefully staged steps, tight control of conditions, and a good deal of patience. Yet, a bacterial enzyme can do it in a single pass, in water, at room temperature, getting every three-dimensional detail right. It would be a great step forward if we could borrow some of these tricks for ourselves.”
Enzymes in this family work like a factory assembly line. As they pass an unfinished molecule from one station to the next, their parts swing around considerably. However, this motion makes them hard to photograph. When several are frozen in motion, each molecule gets caught in a slightly different pose, blurring the averaged image. Moreover, this enzyme is also prone to clumping together in solution.
The researchers approached the problem from two sides, chemistry and structural biology, simultaneously. With their “chemist” hats on, they designed and synthesized small molecules that imitated the unfinished intermediates the enzyme made during its cascade reaction. On the structural biology side, they searched for a way to stabilize the enzyme and keep it from clumping together.
“For a long time, the project seemed impossible,” Tanifuji recalls. “The enzyme simply would not behave in solution, and without a well-dispersed sample there was nothing to image. The turning point was finding a small partner protein called SacE, coded within the same stretch of bacterial genome, which dramatically improved the behavior of the enzyme in solution.”
With both problems solved, the scientists could then flash-freeze these samples in a thin layer of ice and image them using a cryo-electron microscope at Tohoku University. The resolution of the three-dimensional model, reconstructed from thousands of individual enzyme images, was high enough to reveal molecular details at near-atomic level.
“Our structure captures an early moment in the enzyme cycle, the point where the building block has just been loaded. The later steps, where the rings are closed and the finished product is released, are still out of focus. Capturing those states is our next target, and we expect it will take both better probes and new ways of sorting the images,” Tanifuji says, looking to the future.
Journal of the American Chemical Society
Imaging analysis
Not applicable
Cryo-EM Structure of a Monomeric Safracin B Nonribosomal Peptide Synthetase Enabled by an MbtH-Like Protein and Covalent Probe Trapping
19-Oct-2026