Ladderane lipids, with a ladder-like structure of fused cyclobutane rings in their hydrophobic tails, are exclusively found in bacteria that perform anaerobic ammonia oxidation (anammox), suggesting that they play a crucial role in anammox metabolism. As anammox bacteria have not been grown in pure culture and grow extremely slowly in enrichment cultures, researchers have not been able to isolate sufficient quantities of pure ladderane lipids to measure the biophysical properties of ladderane membranes. Without knowledge of the physical properties of ladderane lipids or tools for studying the lipids <i>in vivo</i>, their biological functions and potential applications remain unknown. To answer these questions, we have developed efficient total syntheses of naturally occurring ladderane phospholipids and unnatural analogs for full structure-function studies. We show that ladderane lipids have physical properties that are distinct from conventional straight-chain lipids. Ladderane lipids form dense bilayers with slow lateral diffusion and dense monolayers with low compressibility. These physical properties result in membranes with rates of transbilayer proton diffusion that are an order of magnitude slower than for membranes composed of straight-chain phospholipids. As a proton gradient is used to drive ATP synthesis in anammox bacteria, these results suggest that ladderane lipids in the anammoxosome may prevent the dissipation of the proton gradient during the slow anammox metabolism. To explore how ladderane lipids form such impermeable membranes, we use Nile red solvatochromism to show that the hydrophobic interior of ladderane bilayers is highly nonpolar, likely due to a smaller number of water molecules in the bilayer interior. These results provide experimental evidence for the theoretical model of membrane permeability that involves proton conduction through water wires. On the other hand, small ions and molecules appear to diffuse across membranes via a direct partitioning/diffusion mechanism. The synthesis of ladderane lipids has increased the chemical diversity of hydrophobic tail structure in phospholipids available to explore the structure and function of lipid membranes, including ion permeation. Additionally, the physical properties of ladderane membranes may make them useful for reconstituting membrane proteins, creating stable membrane-based sensors, and drug delivery.

Ladderane phospholipids form dense, low-polarity membranes with low proton/hydroxide permeability

256th National Meeting (2018)

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