CryoPRISM: Imaging Biomolecules in Their Natural Context

Structural biologists traditionally face significant hurdles in studying biomolecular complexes. Extracting these complexes for better imaging can inadvertently alter their natural appearance, while conducting in-situ studies within their native environment remains technically challenging.

To address this critical need, a groundbreaking new method named purification-free ribosome imaging from subcellular mixtures (cryoPRISM) has been developed. Created by Mira May and Gabriela López-Pérez in the Davis lab at MIT, cryoPRISM offers a novel approach to visualize molecular complexes.

cryoPRISM allows visualization of molecular complexes with minimal disruption to their natural context.

Method Overview

cryoPRISM operates by capturing molecular structures from cells that have just been broken open. This ingenious approach is designed to preserve native cellular contacts while still achieving high-resolution molecular details. Associate professor of biology Joey Davis, a lead on the study, highlighted the method's potential, noting that even in well-studied systems like translation in E. coli, new states are continually being discovered.

The Genesis of cryoPRISM

The development of cryoPRISM originated from an unexpected yet pivotal observation made by Mira May, a co-first author of the study. While engaged in a different project aimed at identifying new players in ribosomal regulation using cryoEM, May included unpurified bacterial lysate as a negative control. Contrary to her initial expectations, this control sample yielded clear images of intact ribosomes along with their natural interacting partners. This serendipitous discovery laid the foundation for cryoPRISM.

Key Discoveries and Evolutionary Insights

Using the cryoPRISM method, researchers have already made significant strides, identifying novel ribosomal biology, not just recapitulating previously observed states. One particularly significant finding sheds new light on the evolution of translation regulation.

During periods of environmental stress or unfavorable conditions, such as colder temperatures, bacterial cells instinctively reduce protein translation, leading to a state where ribosomes become idle. These inactive ribosomes are typically blocked by a hibernation factor known as RaiA to prevent their premature reactivation.

May's observations revealed a surprising interaction: some inactive ribosomes were interacting not only with RaiA but also with an elongation factor, EF-G. This was a remarkable discovery because, in bacteria, EF-G was previously thought to interact exclusively with active ribosomes.

This observation suggests that this phenomenon, previously noted only in more complex organisms, may in fact have an older evolutionary origin. This newly identified interaction aligns with a compelling model: elongation factors bind to hibernating ribosomes to protect both the ribosome and the factors themselves from degradation during stress, effectively functioning as a form of short-term storage until conditions improve.

Future Directions and Broader Impact

The utility of cryoPRISM is already expanding. The method is currently being applied to visualize ribosomes in difficult-to-study cells, including pathogenic organisms and red blood cells from patients. cryoPRISM represents a significant step towards the broader goal of structural biology to study biomolecules within their natural cellular environment, offering a powerful new tool for understanding life's fundamental processes.