Phosphorylation, oligomerization and self-assembly in water under potential prebiotic conditions

Abstract

Prebiotic phosphorylation of (pre)biological substrates under aqueous conditions is a critical step in the origins of life. Previous investigations have had limited success and/or require unique environments that are incompatible with subsequent generation of the corresponding oligomers or higher-order structures. Here, we demonstrate that diamidophosphate (DAP)-a plausible prebiotic agent produced from trimetaphosphate-efficiently (amido)phosphorylates a wide variety of (pre)biological building blocks (nucleosides/tides, amino acids and lipid precursors) under aqueous (solution/paste) conditions, without the need for a condensing agent. Significantly, higher-order structures (oligonucleotides, peptides and liposomes) are formed under the same phosphorylation reaction conditions. This plausible prebiotic phosphorylation process under similar reaction conditions could enable the systems chemistry of the three classes of (pre)biologically relevant molecules and their oligomers, in a single-pot aqueous environment.

Figure 1:. DAP mediated phosphorylation of nucleosides/tides demonstrating the potential to generate and progress through the successive levels of nucleotides and oligonucleotides under similar conditions.

Phosphorylation of (a) nucleosides, and 2’- and 3’-mononucleotides to give cyclophosphates, (b) uridine under paste-reaction conditions forming also oligonucleotides, (c) 5’-nucleotides, and (d) 5’- and 3’-phosphorylated oligonucleotides by diamidophosphate (DAP). The reactions were carried out by method A (aqueous, pH 5.5), by method B (aqueous, pH ≈ 7) with or without Mg²⁺ or Zn²⁺ or imidazole, or by method C (paste-reaction). For details see supplementary information and supplementary Tables 1–11.

Figure 2:. DAP induced phosphorylation and esterification of glycerol with short chain fatty acids gives rise to simple mimics of phospholipids leading to formation of protocell like structures under the same reaction conditions signifying the single pot transition of simple building blocks to higher order self-assemblies.

Phosphorylation reaction of DAP (a) with glycerol, and (b) with nonanoic acid and glycerol under the paste-reaction conditions. (c) Synthesis of authentic cyclophospholipid, 26. (d) Transmission electron microscope (TEM) image of a sample (15 mg in 1 mL water) of crude reaction from Fig. 2b showing the formation of vesicle-like structures with a diameter ca. 9.2 μm. (e) TEM image of a sample (1 mg in 1 mL water) of authentic phospholipid 26 from Fig. 2c showing the formation of vesicle-like structures with a diameter ca. 3.5 μm. (F-H) Confocal laser scanning microscopy fluorescence images of vesicles prepared with authentic phospholipid 26 (1 mg in 0.1 mL water) with dye-encapsulation. (f) Green fluorescence indicates hydrophilic pyranine dye encapsulated within the cavity of the liposome. (g) Red fluorescence indicates rhodamine b dye labelling the bilayer phospholipid membrane of the liposome. (h) Fluorescence merged image of phospholipid vesicle prepared with both rhodamine b dye and pyranine dye. For details see supplementary information.

Figure 3:. DAP phosphorylates amino acids in water while activating them towards formation of oligopeptides in the same reaction setting.

(a) Reaction of DAP with glycine, aspartate or glutamate leading to oligopeptides. (b) Reaction of glycine with DAP leading to (quantitatively) phosphorylated species which are intermediates on the way to oligopeptide formation, as seen by ¹H-NMR (b₁), ¹³C-NMR (b₂-b₃) and ¹H-coupled-³¹P-NMR (b₄-b₆) of the reaction mixture. For conditions and details see supplementary information.