Proline and Isoleucine: Indicators of Metabolic Flux and NADPH Balance

Hydrogen isotope ratios (δ²H) of plants and animals are largely determined by the H in water for autotrophs and a combination of H in water and food in heterotrophs. On a finer scale, the δ²H of individual amino acids within a single tissue or organism display extreme variation (~200-800 ‰), with proline and isoleucine typically having the most positive and negative values, respectively. Both of these amino acids involve NADPH in their synthesis pathways, therefore have the potential to reflect the H flux and balance of NADPH (Wijker et al. 2019). Across taxa, the flux of H and the balance of NADP+ and NADPH are dependent on physiological parameters like growth rate, as well as diet quality. In plants, NADPH is generated primarily through photosystem I, but later modified by enzymes involved in secondary metabolisms, including pathways for amino acid synthesis. In heterotrophic organisms, NADPH is generated via transhydrogenases and dehydrogenases. Here we relate the δ²H of proline to modeled metabolic flux rates in organisms as diverse as plants, microbes, fish, mammals, and marine and terrestrial birds by presenting novel amino acid δ²H data. These results should be illustrative in expanding existing theory (Wijker et al. 2019) to include predictions for amino acid metabolism and how it relates to redox balances. Proline δ²H data will be compared to those of isoleucine, an amino acid that is essential in all higher organisms and readily incorporated through direct routing from complex diets. δ²H data will be supported by amino acid carbon (δ¹³C) and nitrogen (δ¹⁵N) isotope data, which has the potential to illuminate whether or not proline is an essential amino acid or synthesized by the organism. The δ²H of proline also shows promise of becoming a new tracer of trophic level, comparable to the d¹⁵N of certain amino acids.

Wijker, R. S. et al., PNAS 116: 12173-12182.