From Amino Acids to α-Keto Acids via β-Elimination and Transamination Initiates a Pathway to Prebiotic Reaction Networks

Abstract

α-Keto acids, such as pyruvate and glyoxylate, may have been critical in generating reaction networks at the origins of life due to their facile carbon-carbon bond formation and their hydrolytic stability. However, demonstrated prebiotic sources of these small α-keto acids have been limited by conditions required for their production, which are not conducive for subsequent incorporation or transition into (proto)metabolic pathways. Here, we demonstrate an abiotic generation of α-keto acids from only two amino acids, starting with the phosphorylation and dehydration of serine (Ser) coupled with a transamination with glycine (Gly), to produce both pyruvate and glyoxylate. This triggers an in situ reaction pathway producing higher-order α-keto acids, including amino acid precursors found in modern biology. These findings may help elucidate how protometabolic chemical networks can emerge on the early Earth under mild aqueous conditions, leading to a coupled amino acid-α-keto acid chemical system capable of supporting a more robust metabolism.

Keywords: Origins of life; Phosphorylation; Phosphoserine; Protometabolism; Pyruvate.

Scheme 1

Routes to α‐keto acids from amino acids. a) The conversion of an α‐amino acid to an α‐keto acid via transamination requires an additional α‐keto acid; however, b) the β‐elimination of a side‐chain activated amino acid produces an α‐keto acid (pyruvate) directly.

Scheme 2

The phosphorylation and β‐elimination of Ser, coupled with a transamination with Gly. This series of reactions yields the two simplest α‐keto acids, pyruvate, and glyoxylate which undergo aldol and Cannizzaro chemistry initiating a reaction sequence generating higher order α‐keto acids, including TCA cycle intermediates.

Figure 1

Ser phosphorylation with DAP. a) Reaction scheme showing the conversion of Ser to PSer in the presence of DAP. ¹H NMR spectra of dry solids redissolved in D₂O from: b) cycle 1 (measured pD = 9.0), c) cycle 2 (measured pD = 8.4), and d) cycle 3 (measured pD = 7.4). e) ³¹P NMR spectrum of cycle‐3 redissolved solids. *PSer conversion after cycle 3 (61%) was determined by ¹H NMR integration. For full ¹H spectra see Figure S18; also see Figure S21.

Figure 2

The β‐elimination of PSer to pyruvate. a) Reaction scheme showing the conversion of PSer to pyruvate via a dehydroalanine intermediate. ¹H NMR spectra of: b) PSer at reaction start, c) 200 mM PSer in 0.5 M phosphate buffer, pH 5.7, heated to 85 °C for 3 days yields 20% pyruvate, d) 200 mM PSer in water, pH 6.5, heated to 85 °C for 3 h yields 0.2% pyruvate, and e) 200 mM PSer in water, pH 6.5, heated to 85 °C for 3 h in the presence of 0.1 equiv. CuCl₂ yields 72% pyruvate (68%–77% in quadruplicate). Note: The 4.8 ppm region (HOD/H₂O peak) is removed to enhance clarity. *Pyruvate (keto + hydrate) yields were calculated using a 100 µL reaction aliquot diluted to 600 µL with D₂O containing t‐butanol as an internal standard.

Scheme 3

Copper chelation of PSer catalyzes the β‐elimination of phosphate. One of the three rotamers (left) of the copper‐chelated phosphoserine has three chelation sites and sets an anti‐conformation (H in red), ideal for a concerted β‐elimination. Copper complex representations are here simplified, not showing the water ligands.

Figure 3

Generation of α‐ketoglutarate from PSer and Gly. a) Reaction scheme showing the production of α‐ketoglutarate from pyruvate (via β‐elimination of PSer) and glyoxylate (via transamination with Gly and pyruvate). b) ¹H NMR spectrum of 200 mM PSer, 1 equiv. Gly, 0.1 equiv. CuCl₂, in water at pH 5 heated at 96 °C for 24 h. For full spectrum, see Figure S31. c) ¹³C NMR spectrum of 200 mM PSer, 1 equiv. ¹³C(2)‐Gly, 0.1 equiv. CuCl₂, in water at pH 5 heated at 96 °C for 24 h.

Scheme 4

¹³C‐labeled studies. ¹³C(2)‐labeled Gly generates ¹³C(2)‐labeled glyoxylate, resulting in a distinct labeling pattern for downstream products including maloyl formate, α‐ketoglutarate, and aconitoyl formate. Asterisks mark ¹³C‐labeled positions above. The use of ¹³C(1,2)‐dilabeled Gly also yields labeling patterns consistent with the reaction scheme (Figures S10–S16).