Molecular scissors caught in action: A structural blueprint for RNA therapeutics

LA JOLLA, CA-RNA interference is a natural mechanism for living cells to control whether specific genes are being used or not.

Crowned with the 2006 Nobel Prize for Physiology and Medicine, the discovery of RNA interference has since been harnessed by scientists to create a powerful and growing class of drugs capable of suppressing disease-related genes. Seven such drugs have already received FDA approval, including inclisiran, which can replace daily cholesterol-lowering pills with biannual injections. Despite these clinical successes, the molecular details of how the system executes its cuts remained poorly understood.

Now, Scripps Research scientists have captured the first high-resolution structural images of the human RNA interference molecular machinery in its slicing-ready state. Their findings, published in Nature Structural & Molecular Biology on June 24, 2026, identify the precise atomic interactions that determine when and where the machinery cuts. The structures show which building blocks of the protein are key to its function and provide a mechanistic explanation for how some RNA sequences are better than others at cutting their targets.

Cryo-EM structure of human Argonaute 2 bound to a paired guide-target RNA in a catalytic state. Credit: Scripps Research

Click here for a high-resolution image. RNA interference has become a powerful platform for treating disease, with more drugs entering the pipeline every year, says Scripps Research professor Ian MacRae, the senior author of the new study. But until now, we've been designing these drugs somewhat in the dark. This work finally gives us the structural picture we need to understand what makes a good therapeutic siRNA and how to design better ones.

The challenge for drug developers is that, for any given disease-causing gene, thousands of different siRNA sequences could potentially shut down a corresponding target RNA. Researchers have been unable to predict which ones succeed without testing them, leading to a long trial-and-error process to develop new RNA interference drugs.

In the new work, postdoctoral associate Sucharita Sarkar and staff scientist Luca Gebert, co-first authors from MacRae's laboratory, set out to determine the high-resolution structure of Argonaute 2 in a cutting-ready state-bound to an siRNA molecule and poised to cut the corresponding messenger RNA. Their challenge was that this state only lasts a fleeting moment.

The trick was that we had to come up with a special set of mutations that would stabilize this active conformation, says Gebert.

Once they identified these mutations, the team used cryo-electron microscopy (cryo-EM) to capture the atomic-level arrangement of Argonaute 2 in the moments just before it cuts its RNA target. That structure revealed something surprising: Rather than sitting in a straight, natural conformation inside Argonaute 2, the guide-targeted RNA duplex was physically distorted. This coordinated deformation, imparted by Argonaute 2, positions the precise chemical bond that must be cleaved directly within the protein's molecular scissors.

For the first time, we could see exactly where two previously overlooked amino acids sit in the active site-and their positions redefine how we understand Argonaute 2 catalysis, says Sarkar.

Those two amino acids-Lysine709 and Arginine710-help drive the cutting reaction, working alongside the four other amino acids already known to be important for cutting RNA. Lysine709 acts as a molecular checkpoint, held away from the active site until extended guide-target pairing triggers the duplex deformation that releases it into cutting position. Arginine710 fine-tunes catalytic efficiency by sensing the identity of a specific position in the target RNA, explaining a long-standing but poorly understood empirical rule in siRNA design.

Beyond explaining a key step in RNA interference, the new study could offer specific guidance for how to design siRNA molecules that are most likely to be effective. Sequences and chemical modifications that allow the paired RNA to more readily adopt the distorted shape are predicted to favor activation by Argonaute 2, while those that rigidify the central region are expected to impair it.

MacRae says the work opens the door to what is known as rational design -engineering siRNA sequences based on structural principles rather than trial and error.

This class of drug is already impressive, and the drugs in development are only getting more powerful, he says. Understanding the mechanism at this level should allow us to design better drugs from the start, which could expand the range of diseases we're able to treat with RNA interference.

The study Catalytic activation of human Argonaute 2 requires RNA duplex deformation was authored by Sucharita Sarkar, Luca F. R. Gebert and Ian J. MacRae, all of Scripps Research.

This work was supported by funding from the National Institutes of Health (R35GM127090) and through a sponsored research agreement with Eli Lilly and Company.

LINK:	https://www.scripps.edu/news-events/news/20260624-macrae-molecular-sci...
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