A bioenergetic basis for membrane divergence in archaea and bacteria

Abstract

Membrane bioenergetics are universal, yet the phospholipid membranes of archaea and bacteria-the deepest branches in the tree of life-are fundamentally different. This deep divergence in membrane chemistry is reflected in other stark differences between the two domains, including ion pumping and DNA replication. We resolve this paradox by considering the energy requirements of the last universal common ancestor (LUCA). We develop a mathematical model based on the premise that LUCA depended on natural proton gradients. Our analysis shows that such gradients can power carbon and energy metabolism, but only in leaky cells with a proton permeability equivalent to fatty acid vesicles. Membranes with lower permeability (equivalent to modern phospholipids) collapse free-energy availability, precluding exploitation of natural gradients. Pumping protons across leaky membranes offers no advantage, even when permeability is decreased 1,000-fold. We hypothesize that a sodium-proton antiporter (SPAP) provided the first step towards modern membranes. SPAP increases the free energy available from natural proton gradients by ∼60%, enabling survival in 50-fold lower gradients, thereby facilitating ecological spread and divergence. Critically, SPAP also provides a steadily amplifying advantage to proton pumping as membrane permeability falls, for the first time favoring the evolution of ion-tight phospholipid membranes. The phospholipids of archaea and bacteria incorporate different stereoisomers of glycerol phosphate. We conclude that the enzymes involved took these alternatives by chance in independent populations that had already evolved distinct ion pumps. Our model offers a quantitatively robust explanation for why membrane bioenergetics are universal, yet ion pumps and phospholipid membranes arose later and independently in separate populations. Our findings elucidate the paradox that archaea and bacteria share DNA transcription, ribosomal translation, and ATP synthase, yet differ in equally fundamental traits that depend on the membrane, including DNA replication.

Figure 1. Membrane lipids of archaea and bacteria.

Archaeal lipids (left) are typically composed of isoprenoid chains linked by ether bonds to an sn-glycerol-1-phosphate (G1P) backbone. The chirality of the two glycerol backbones is fully conserved within each clade not only in structure but in their unrelated synthetic enzymes. Although ether linkages have been observed in bacterial membranes and isoprenoids are common to all three domains, bacterial lipids (right) are typically composed of fatty acids in ester linkage to an sn-glycerol-3-phosphate (G3P) skeleton. Despite widespread horizontal gene transfer, no bacterium has been observed with the archaeal enantiomer, or vice versa .

Figure 2. The model.

A cell with a semi-permeable membrane sits at the interface between an alkaline and an acidic fluid. The fluids are continuously replenished and otherwise separated by an inorganic barrier. Hydroxide ions (OH⁻) can flow into the cell from the alkaline side by simple diffusion across the membrane, with protons (H⁺) entering in a similar manner from the acidic side. Other ions (Na⁺, K⁺, Cl⁻, not shown) diffuse similarly, as a function of their permeability, charge, and respective internal and external concentrations on each side. Inside the protocell, H⁺ and OH⁻ can neutralize into water, or leave towards either side. Internal pH thus depends on the water equilibrium and relative influxes of each ion. A protein capable of exploiting the natural proton gradient sits on the acidic side, allowing energy assimilation via ATP production, or carbon assimilation via CO₂ fixation.

Figure 3. Dynamics of free-energy change (−ΔG) in cells powered by natural proton gradients.

(A) Proton-permeable vesicles (≥10⁻⁴ cm/s) have only a small loss of free-energy compared with an open system (pH gradient 7∶10, 1% ATPase). Reduced membrane permeability (≤10⁻⁴ cm/s), including permeabilities equivalent to modern membranes (<10⁻⁵ cm/s), collapse the gradient within seconds. (B) At low permeability (10⁻⁶ cm/s), −ΔG collapses regardless of gradient size. Within seconds, H⁺ flux through ATPase equilibrates with the acidic fluids. (C) The collapse of −ΔG is more extensive the greater the amount of membrane-bound ATPase, even with a leaky membrane (10⁻³ cm/s). (D) With Ech, the collapse of the natural gradient is similar to that of the ATPase, showing that natural proton gradients can power energy (ATPase) and carbon (Ech) metabolism, given 1%–5% enzyme in membrane. Na⁺ permeability was kept 6 orders of magnitude higher than that of H⁺ throughout all simulations in this and all figures of the article. Except in (B), all results were calculated in a pH gradient 7∶10.

Figure 4. Pumping H⁺ or Na⁺ does not offer a sustained selective advantage.

(A) Pumping H⁺ in a membrane with 1% ATPase causes a sustained loss in −ΔG as membrane permeability decreases with 1% pump. Even with 5% pump, −ΔG does not change over 3 orders of magnitude, and pumping only improves −ΔG near modern membrane permeability (≤10⁻⁵ cm/s). (B) Pumping less-permeable Na⁺ is initially better, adding to the natural gradient, but the early benefit is lost as membranes become tighter, due to the collapse of the natural H⁺ gradient. In the absence of a gradient, pumping both H⁺ (C) and Na⁺ (D) offers a sustained advantage to tightening up membranes, but given a minimal requirement of around 15–20 kJ/mol to power aminoacyl adenylation, the energy attained is not sufficient to power intermediary biochemistry.

Figure 5. SPAP significantly increases free energy.

(A) Because external Na⁺ concentration (0.4 M) is higher than H⁺ concentration (10⁻⁷ M), SPAP initially collapses −ΔG, and it takes minutes for the 1∶1 H⁺∶Na⁺ exchange to increase −ΔG; eventually it renders an increase of ∼60%. (B) The greatest increases are attained in relatively alkaline pH 7∶10 environments, saturating as % surface area rises. Despite equivalent gradient sizes, the absolute difference in H⁺ and OH⁻ concentrations means a 6∶9 gradient gives a lower −ΔG, as the rate of H⁺ influx is greater while neutralizing OH⁻ influx is lower. A 5∶8 gradient undermines −ΔG further, with or without SPAP. (C) SPAP facilitates colonization of environments with weaker proton gradients. 1% SPAP pushes −ΔG above 20 kJ/mol in a 7.5∶10 gradient, whereas 10% SPAP salvages an otherwise unviable 8∶10 gradient. All simulations with 1% promiscuous ATPase, no pump, no Ech, and H⁺ permeability 10⁻³ cm/s.

Figure 6. SPAP gives a sustained benefit to pumping favoring tighter membranes and allowing free living.

(A) The combination of SPAP with 5% H⁺ pump gives a sustained increase in −ΔG as membrane permeability decreases, for the first time favoring the evolution of modern proton-tight phospholipid membranes. In contrast, 1% H⁺ pump gives an initial benefit, but provides insufficient power to sustain −ΔG as the gradient is lost with decreasing permeability. (B) The combination of SPAP with both 1% and 5% Na⁺ pump provides an initial benefit, but neither provides enough power to sustain −ΔG with decreasing permeability. (C) SPAP facilitates colonization of smaller gradients, ultimately making it possible to survive, after the evolution of tight membranes, in the total absence of a gradient (D); cells could not survive without a gradient unless relatively proton-tight membranes were already in place, as −ΔG falls well below the 15–20 kJ/mol threshold upon losing the gradient with a leaky membrane. All simulations assume 1% SPAP. Legend in (B) is common to all panels.

Figure 7. Divergence of archaea and bacteria.

(A) Ions cross the membrane in response to concentration gradients and electrical potential. OH⁻ neutralizes incoming protons. The H⁺ gradient can drive energy metabolism via ATPase, and carbon metabolism via Ech (not shown). (B) SPAP generates a Na⁺ gradient from the H⁺ gradient. As Na⁺ is less permeable than H⁺, SPAP improves coupling, given promiscuity of membrane proteins for H⁺ and Na⁺. (C) Membrane pumps generate gradients by extruding H⁺ or Na⁺ ions. (D) Exploiting natural gradients demands high membrane permeability, but pumping with SPAP drives the evolution of tighter membranes, facilitating colonization of less alkaline environments. (E) Impermeable membranes funnel ion flow through bioenergetic proteins, independent of natural gradients. (F) From bottom up, SPAP favors divergence, selection for active pumping and tighter membranes. Pumping and phospholipid membranes arose independently in archaea and bacteria.