Enzyme-free synthesis of natural phospholipids in water

Abstract

All living organisms synthesize phospholipids as the primary constituent of their cell membranes. While phospholipids can spontaneously self-assemble in water to form membranebound vesicles, their aqueous synthesis requires pre-existing membrane-embedded enzymes. This limitation has led to models in which the first cells used simpler types of membrane building blocks and has hampered integration of phospholipid synthesis into artificial cells. Here we demonstrate that a combination of ion pairing and self-assembly of reactants allows highyielding synthesis of cellular phospholipids in water. Acylation of 2-lysophospholipids using cationic thioesters occurs in mildly alkaline solutions resulting in the formation of cell-like membranes. A variety of membrane-forming natural phospholipids can be synthesized. Membrane formation takes place in water from natural alkaline sources, such as soda lakes and hydrothermal oceanic vents. When formed vesicles are transferred to more acidic solutions, electrochemical proton gradients are spontaneously established and maintained.Main Text: Cellular membranes composed of glycerophospholipids are found in all living organisms (1). Bacterial and eukaryotic membranes consist of diacylphospholipids, in which two ester linkages connect a polar head group to two hydrophobic tails. Cells synthesize diacylphospholipids through enzymatic acylation of lysophospholipids (2). As several enzymes involved in phospholipid biosynthesis must themselves be membrane-bound for enzyme activity, it is unclear if phospholipid membranes could have formed in the absence of advanced enzymatic machinery (3-6). Enzyme-free synthesis of glycerophospholipids can occur using wet-dry cycling (7, 8) and acylation of glycerophosphates can take place in the presence of a large excess of activated long-chain acyl imidazole derivatives, but the reactions require organic co-solvent due to the hydrophobicity of the acylating precursors and result in unnatural phospholipids (9, 10). A high-yielding synthesis of natural phospholipids in water would shed light on the origin and evolution of cellular membranes and open up new routes for lipid synthesis in artificial cells (11,12).

Spontaneous emergence of membrane-forming protoamphiphiles from a lipid–amino acid mixture under wet–dry cycles

“… 15,38,40 This is mainly because they are the major constituents of contemporary cell membranes and have also been shown to get synthesized under certain prebiotically relevant conditions. 41–44 Furthermore, they could readily form stable vesicles under varied experimental conditions, making it easy to monitor the effect of membrane on the reaction of interest. As a first step, we asked whether Gly oligomers (peptides) are produced in the presence of POPC and, if yes, what their overall yield might be.…”

“…This signifies the importance of ester-amide exchange in also facilitating the formation of protoamphiphiles (NAA), in addition to that of protopeptides. Furthermore, there has been increasing evidence supporting the prebiotic synthesis of phospholipids, which suggest their potential availability as a precursor for NAA synthesis on the early Earth [41][42][43][44] . Nonetheless, fatty acids and other single chain amphiphiles are widely considered as components of prebiotic membranes, because of their plausible prebiotic abundance and structural simplicity, whereas phospholipids are typically looked as a product of membrane evolution.…”

“…Furthermore, there has been increasing evidence supporting the prebiotic synthesis of phospholipids, which suggest their potential availability as a precursor for NAA synthesis on the early Earth. 41–44 Nonetheless, fatty acids and other single chain amphiphiles are widely considered as components of prebiotic membranes, because of their plausible prebiotic abundance and structural simplicity, whereas phospholipids are typically looked as a product of membrane evolution. Therefore, we are systematically investigating the possibility of ester–amide exchange between amino acids and other chemically simpler prebiotic amphiphiles.…”

“…Phospholipids such as POPC are often used to study the effect of membrane-forming amphiphiles on prebiotically pertinent processes 15,38,40 . This is mainly because they are the major constituents of contemporary cell membranes and have also been shown to get synthesized under certain prebiotically relevant conditions [41][42][43][44] . Furthermore, they could readily form stable vesicles under varied experimental conditions, making it easy to monitor the effect of membrane on the reaction of interest.…”

A self-templated route to monodisperse complex droplets as artificial extremophile-mimic from coacervate-liposome interplay

“…4h), which consistent with the anticipated role of coacervate nucleus as the buffer. This effect on one hand enabled the generation of a transmembrane pH gradient, which might be involved in the evolution and biosynthesis of prebiotic life 39 . On the other hand, it provided a pH homeostatic environment for biomolecules.…”