PepA: an extracellular manganese oxidase induced by lignin in Pseudomonas putida KT2440

Background: Manganese (Mn) oxides play a key role in the oxidation of recalcitrant organic carbon, including lignin (1), and have also emerged as a potential material in a wide range of technological applications, e. g, energy storage or catalysis. A transcriptomic study aimed to identify lignin-induced genes in the environmental model bacterium Pseudomonas putida KT2440 revealed pepA, encoding an animal-heme peroxidase-like enzyme homologous to certain Mn oxidases (2), as one of the main induced genes.
Objectives: To characterise the mechanisms of secretion and action of PepA in P. putida KT2440.
Methods: A collection of P. putida mutant strains was designed to unequivocally asses the function of pepA and that of the adjacent operon coding for a type-1 secretion system (T1SS). Liquid Chromatography coupled to tandem mass spectrometry (LC-MS/MS) was used to confirm localization and processing of the mature enzyme. Transmission (TEM) and scanning (SEM) electron microscopy techniques were applied to detect and characterise manganese oxides nanoparticles (MnOx NPs). Superoxide generation by PepA was tested by making use of a chemoluminescent probe.
Results: This work shows that PepA is an extracellular heme-dependent enzyme that oxidizes Mn(II) by generating superoxide. A specific T1SS encoded adjacent to pepA is involved in PepA transport and processing of the mature form. A PepA-overproducer P. putida strain allowed the characterisation of the extracellular MnOx NPs generated. These results expand the knowledge on bacterial manganese oxidases and their potential role in lignin depolymerization, and broaden the biotechnological utilities of the model P. putida KT2440 bacterial chassis.