We poorly understand how molecular components of life evolve. In absence of a fossil record, we rely on comparisons between contemporary protein complexes to make inferences about molecular evolution. Bacterial flagellar motors are structurally diverse motility devices, well-suited to evolutionary study. Motors across different lineages evolved varied elaborations on top of a highly conserved mechanistic core. These additional scaffolds make for wider motors, increasing torque output for navigating high-viscosity environments. Variety in presence and composition of scaffolds allow comparison between species, to understand how novel proteins contribute to motility and modify existing complexes.

Towards understanding how new components evolve, I studied flagellar structure and phylogenetics in pathogen Campylobacter jejuni. Its high-torque motor contains an outer membrane-associated basal disk and a periplasmic scaffold. I used cryo-electron tomography in situ imaging to expand the list of proteins that make up the basal disk and periplasmic scaffold. Next, I reconstructed a sub-nanometre resolution cryo-electron microscopy map of the C. jejuni motor. I combined homology and de novo protein modelling to assemble a near-complete atomic model of the motor, building on my previous results. I validated the model by in vitro protein interactions and in situ imaging of deletion mutants. My model revealed how additional motor components assemble to enable high torque, and how pre-existing parts changed to accommodate this. Structures revealed that various components are likely homologous to contemporary enzymes and other non-flagellar proteins. Finally, I conducted a bioinformatics analysis, showing most proteins of the C. jejuni scaffold are unique to species within its phylum, Campylobacterota. I found the major basal disk protein is additionally present in δ-proteobacteria, and described a new closely related protein family. Combining my analysis with published results from our lab, I implicate δ-proteobacteria as possibly descendant from an ancestral motor before their divergence from Campylobacterota.