Collaborative Research: RUI: Structure-Function Relationships and Efficiency of Bacterial Flagellar Motors Using Computational Fluid Dynamics and Directed Evolution Experiments

Centre College Of Kentucky, Danville KY

Investigators

Abstract

Bacteria are among the oldest organisms on Earth. Through biological evolution, many bacteria have acquired a biomechanical flagellum consisting of a helical appendage that is rotated by a molecular motor to move them through their fluid environment. Flagellar genetic organization, regulation, and structure are broadly conserved across bacterial species, and flagella play vital roles in the bacterial life cycle, including in host-microbe interactions. Thus, understanding bacterial motility has broad implications in medicine, in biology, and in the development of micro-robotic swimmers. This research aims to understand how the energy efficiency of the bacterial motor has influenced the evolutionary development of the bacterial motility system. The project will create precisely calibrated computational tools to study variants of the medically important organism Pseudomonas aeruginosa with different motor properties. The energy efficiency of each variant will be measured and associated with structural changes in motor components. The computational tools and a library of these motor variants will be made publicly available so that other researchers may use them for related research. The experiments and computations required for this work will all be conducted in tandem with undergraduate students, with the goal that many of them, including students from underrepresented backgrounds, will pursue careers in interdisciplinary scientific research. Recent work has uncovered the molecular structure of the stator unit responsible for generating torque in the bacterial flagellar motor. This project aims to characterize the relationships between specific structural properties of the bacterial stator and its energy efficiency. Strains of P. aeruginosa with suboptimal stators will be created and used in directed evolution experiments to select for variant strains with improved motility. Quantitative microscopy will be used to measure bacterial motion as they move through different fluid environments. Experimentally determined trajectories will be input into precisely calibrated computational fluid dynamics simulations to determine the energy efficiency of the stator. The determination of sequence and structural features of evolved stators with different energy efficiencies will provide insights into the relationship between the mechanical properties and biomolecular structures of nanomotors. Additionally, to study how the presence of a nearby boundary affects the swimming efficiency of the evolved strains, motility experiments will be performed near smooth surfaces and surfaces with tunable viscosity and electrostatic properties. The surfaces will be constructed from lipids and proteins to mimic the environments that P. aeruginosa commonly encounters in its natural environment as it undergoes motile-to-sessile transitions. The calibration measurements will allow other researchers to create additional precisely calibrated computational fluid dynamics models and develop similar computational probes as used in this project. The publicly available library of evolved motor variants and the structure-function map for the stator will deepen the understanding of the relationship between motor proteins and motor performance. This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.