Compartments, Membranes and Transporters: A Computational Investigation.

Abstract

Compartmentalisation is fundamental to cellular life, necessitating both a barrier, most often a membrane, and associated transport machinery that controls the influx and efflux of critical substrates. The dynamics and behaviour of the diverse macromolecular assemblies that are compartments, membranes, and their transporters are critical in a range of cellular processes across all domains of life. Computational chemistry techniques are uniquely placed to provide insight into these processes that are often difficult to probe with experimental tools. In this thesis, computational chemistry is used to provide insight into the dynamics of compartments, membranes, and transporters with the aim of exploring their complexities. The link between lipid structural diversity and membrane dynamics in eukaryotic plasma membranes was explored, demonstrating the multilevel influence of the membrane composition on resultant biophysical properties. Firstly, the link between lipid diversity of the neuronal plasma membrane and its biophysical properties was investigated, finding that headgroup diversity, and not acyl-tail diversity, was the primary driver of variation in membrane biophysical properties. Further, a complex membrane model for the epithelial plasma membrane was developed and its biophysical properties related to its unique composition, including plasmalogens, Forssman lipids, and hydroxylated sphingolipids uncommon in other membranes. The role of the membrane in the interplay between pathogen and host was explored, highlighting the role of host-derived lipids in modulating bacterial membrane biophysical properties and transporter function. Firstly, a complex membrane model for the Acinetobacter baumannii inner membrane in the presence and absence of host-derived polyunsaturated lipids was constructed. Incorporation of host-derived lipids resulted in dramatic changes to membrane biophysical properties. Further, the effect of these changes on the function of the AdeABC and AdeIJK multidrug efflux pumps was examined, finding that incorporation of host-derived lipids can modulate AdeABC dynamics and efflux, linking membrane composition to the evolution of antimicrobial resistance. The mechanisms of transmembrane and trans-compartment ion transport were explored. Firstly, the dynamics of the PsaBCA bacterial Mn2+ import system were investigated. Determinants of Mn2+ specificity in the substrate binding protein PsaA were examined, finding that water-labile metal coordination is critical for selectivity. Secondly, the dynamics of an inward-facing structure of the cognate PsaBC Mn2+ importer were determined, identifying a metal coordination site in the permease domain absent in structures of other type II ATP binding cassette (ABC) importers. Additionally, determinants of metal ion transport and selectivity in a series of engineered bacterial protein cages was investigated, with rational design of pore mutants modulating pore dynamics and ion flux. Finally, the mechanism of action of a novel series of proton transporting mitochondrial uncouplers was determined and their structure-activity relationship explored. Overall, this thesis illuminates the complexities of compartments, membranes, and transporters, highlights their central role in the fundamental processes of life, and explores the integrated approaches required to understand them.