Prebiotic cationic amino acids support formation of an ancient protein fold

Abstract

The evolutionary basis for the selection of the standard proteinogenic amino acids remains elusive, particularly for the long cationic amino acids, lysine and arginine, which were likely scarce in the prebiotic environment. In contrast, shorter cationic amino acids, such as ornithine (Orn), and 2,4-diaminobutyric acid (Dab), are thought to have been more abundant. Here, we investigated whether these prebiotic cationic amino acids can support protein tertiary structure formation, using computational protein design, biophysical and crystallographic analyses, and molecular dynamics (MD) simulations. We designed sequences for an ancient protein fold, the double-{Psi} {beta}-barrel (DPBB), with ornithine and plausible prebiotic amino acid sets. Although all the designed sequences were unfolded under standard dilute aqueous conditions, one Orn-containing variant folded in highly concentrated conditions. Crystallographic analysis revealed that both this peptide and its Dab-substituted derivative adopted the double-Z {beta}-barrel (DZBB) fold, a likely evolutionary intermediate between extant {beta}-barrel folds. Therefore, Orn and Dab might have supported the foldability of primitive proteins before the incorporation of lysine and arginine into the genetic code.