Molecular model of a bacterial flagellar motorin situreveals a “parts-list” of protein adaptations to increase torque

Abstract

One hurdle to understanding how molecular machines function and evolve is our inability to see their structuresin situ. Here we describe a minicell system that enablesin situcryogenic electron microscopy imaging and single particle analysis to probe the mechanisms and evolution of an iconic molecular machine, the bacterial flagellar motor, which spins a helical propeller for bacterial propulsion. Innovations in sample preparation and imaging enabled resolutions sufficient to build anin situmolecular model of theC. jejuniflagellar motor. Our results provide unprecedented insights into thein situcontext of flagellar motors, highlight origins of recruited components involved in the unusually high torque of theC. jejunimotor, identify previously unknown components, and reveal corresponding modifications of core components. We also visualise structures involved in torque generation and secretion previously recalcitrant to structure determination. This technique will be of broad applicability to other large membrane-residing protein complexes. Note that this manuscript has a sibling manuscript titled “Evolution of a large periplasmic disk in Campylobacterota flagella facilitated efficient motility alongside autoagglutination” that dissects the function of the large disk described in this manuscript.