Anoxic metabolism and biochemical production in Pseudomonas putida F1 driven by a bioelectrochemical system

Abstract

BackgroundPseudomonas putida is a promising host for the bioproduction of chemicals, but its industrial applications are significantly limited by its obligate aerobic character. The aim of this paper is to empower the anoxic metabolism of wild-type Pseudomonas putida to enable bioproduction anaerobically, with the redox power from a bioelectrochemical system (BES).ResultsThe obligate aerobe Pseudomonas putida F1 was able to survive and produce almost exclusively 2–Keto-gluconate from glucose under anoxic conditions due to redox balancing with electron mediators in a BES. 2-Keto-gluconate, a precursor for industrial anti-oxidant production, was produced at an overall carbon yield of over 90 % based on glucose. Seven different mediator compounds were tested, and only those with redox potential above 0.207 V (vs standard hydrogen electrode) showed interaction with the cells. The productivity increased with the increasing redox potential of the mediator, indicating this was a key factor affecting the anoxic production process. P. putida cells survived under anaerobic conditions, and limited biofilm formation could be observed on the anode’s surface. Analysis of the intracellular pools of ATP, ADP and AMP showed that cells had an increased adenylate energy charge suggesting that cells were able to generate energy using the anode as terminal electron acceptor. The analysis of NAD(H) and NADP(H) showed that in the presence of specific extracellular electron acceptors, the NADP(H) pool was more oxidised, while the NAD(H) pool was unchanged. This implies a growth limitation under anaerobic conditions due to a shortage of NADPH and provides a way to limit biomass formation, while allowing cell maintenance and catalysis at high purity and yield.ConclusionsFor the first time, this study proved the principle that a BES-driven bioconversion of glucose can be achieved for a wild-type obligate aerobe. This non-growth bioconversion was in high yields, high purity and also could deliver the necessary metabolic energy for cell maintenance. By combining this approach with metabolic engineering strategies, this could prove to be a powerful new way to produce bio-chemicals and fuels from renewables in both high yield and high purity.Electronic supplementary materialThe online version of this article (doi:10.1186/s13068-016-0452-y) contains supplementary material, which is available to authorized users.

Anodic respiration of Pseudomonas putida KT2440 in a stirred-tank bioreactor

“…In consistence with published results with P. putida F1 [25], the current observed with P. putida KT2440 and riboflavin was negligibly small compared to the current observed with K 3 Fe(CN) 6 (data not shown).…”

“…With citrate as substrate pHBA formation was possible. In anodic electrofermentations with P. putida F1 it had been shown that during the anodic oxidation of glucose with P. putida F1 a NADPH shortage occurred [25]. Because P. putida F1 and P.…”

Resource Recovery from Wastewater by Biological Technologies: Opportunities, Challenges, and Prospects

“…On the side of the anode, electricity production dominated the focus so far, but in a recent study it has been shown that using a pure culture of a strict aerobe, breathing the anode allowed the high yield production of a chemical precursor for industrial antioxidant production (Lai et al, 2016). This is a key development, as it highlights the advantage of using a strict aerobe in such a system.…”

Metabolic engineering and bio-electrochemical synthesis in Pseudomonas putida KT2440 for the production of p-hydroxybenzoate and 2-ketogluconate

“…Its physiological and metabolic robustness highlights its advantages in biotechnology and environmental applications (Fernández et al, 2012b;Verhoef et al, 2009;Wierckx et al, 2005;Wittgens et al, 2011), and also provokes an increasing interest in developing electrochemical production processes Lai et al, 2016;Schmitz et al, 2015). Recently, Pseudomonas putida was recommended as the next generation synthetic biology chassis (Adams, 2016).…”

“…The modification and optimization of microbes for the target products biosynthesis in electrochemical system, is progressing slowly mainly due to the little scientific insight of electron transfer. Low conversion rate and productivity can be seen from many reported cases Jeon et al, 2015;Lai et al, 2016;Nevin et al, 2010), highlighting the utmost necessity to improve the electron transfer rate.…”