Rationally Engineering pH Adaptation of Acid-Induced Arginine Decarboxylase from Escherichia coli to Alkaline Environments to Efficiently Biosynthesize Putrescine

Abstract

Acid-induced arginine decarboxylase AdiA is a typical homo-oligomeric protein biosynthesizing alkaline nylon monomer putrescine. However, upon loss of the AdiA decamer oligomeric state at neutral and alkaline conditions the activity also diminishes, obstructing the whole-cell biosynthesis of alkaline putrescine. Here, a structure cohesion strategy is proposed to change the pH adaptation of AdiA to alkaline environments based on the rational engineering of meridional and latitudinal oligomerization interfaces. After integrating substitutions of E467K at the latitudinal interface and H736E at the meridional channel interface, the structural stability of AdiA decamer and its substrate transport efficiency at neutral and alkaline conditions are improved. Finally, E467K_H736E is well adapted to neutral and alkaline environments (pH 7.0-9.0), and its enzymatic activity is 35-fold higher than that of wild AdiA at pH 8.0. Using E467K_H736E in the putrescine synthesis pathway, the titer of putrescine is up to 128.9 g·L^-1 with a conversion of 0.94 mol·mol^-1 in whole-cell catalysis. Additionally, the neutral pH adaptation of lysine decarboxylase, with a decamer structure similar to AdiA, is also improved using this cohesion strategy, providing an option for pH-adaptation engineering of other oligomeric decarboxylases.

Keywords: arginine decarboxylase; biocatalysis; pH adaptation; protein engineering; putrescine.