Interior of one of Michigan State's cryogenic electron microscopes. Credit: Paul Henderson / Michigan State University College of Natural Science.
Researchers at Michigan State University have captured, for the first time, high-resolution experimentally determined structures of SpoIVFB, an elusive protein.
Found in the model bacterium Bacillus subtilis, SpoIVFB belongs to a family of specialized enzymes known as intramembrane proteases. These enzymes help regulate key cellular functions across all kingdoms of life. Notably, SpoIVFB plays a critical role in sporulation—the process that allows bacteria to survive harsh conditions such as radiation and heat.
In the years-long quest to characterize SpoIVFB, researchers had traditionally used X-ray crystallography. The quality of SpoIVFB’s crystals and the quantity of sample required, however, were two consistent hurdles. So, they turned to cryo-EM.
Cryo-EM was able to overcome X-ray crystallography’s biggest bottlenecks and visualize SpoIVFB like never before.
Through cryo-EM imaging and molecular dynamics, researchers Lee Kroos and Ben Orlando confirmed the mechanism SpoIVFB uses to prepare its substrate for cleaving, as well as the route water might take to reach this site. The cryo-EM structures determined by Orlando and Kroos demonstrate that SpoIVFB engages the substrate through an interaction known as beta-sheet augmentation.
This revelation was the final piece of a vast biological puzzle. Four classes of intramembrane proteases are found across the kingdoms of life, from the simplest bacterium to humans. Three of these were known to engage their substrates through beta-sheet augmentation.
This study offers insight into a mechanism of cellular regulation found in organisms ranging from bacteria to humans, with vast implications for microbiology, structural biology, enzymology and human disease.
Information from Michigan State University