(175af) Continuous Production of 1.3-Propanediol By Metabolically Engineered Cyanobacteria Employing an Airlift Photobioreactor with pH-Stat System

The 4th generation biorefinery research using recombinant microalgae is ongoing in many countries because the direct production of fuel/chemicals from carbon dioxide by photogenic microorganisms is essentially sustainable. Among various photosynthetic microorganisms, cyanobacteria has been recognized as proper platform of a metabolic engineering for chemicals production due to the well-established genetic tools as well as its high growth rate. In our previous research, we developed a genetically engineered Synechococcus elongatus producing 1,3-propanediol (1,3-PDO), which is an important chemical used for the synthesis of polytrimethylene terephthalate (PTT) (1, 2). In order to achieve the cost-effective production of 1,3-PDO using the genetically engineered S. elongatus, it is important to develop an efficient continuous photobioreactor system (PBS). Among various types of PBSs, it is known that an airlift bioreactor offers high mass transfer rate for carbon dioxide dissolution, less share stress against algae and low operational power consumption for mixing. In this study, we examined the possibity of applying an airlift photobioreactor (ALPB) for the cultivation of cyanobacteria and performed the continuous production of 1,3-PDO by the airlift photobioreactor with pH-stat system.

A genetically engineered S. elongatus producing 1,3-propanediol (1,3-PDO) (referred to as TA2984) was constructed by introducing a metabolic pathway to synthesize 1,3-PDO from DHAP via glycerol into S. elongatus PCC7942 by homologous recombination (1). A glass-made airlift photobioreactor system with an internal draft tube in the central section of the reactor was employed for the cultivation of the strain for 1,3-PDO production.

Firstly, batch experiments employing TA2984 were conducted by the use of ALPB. The results showed that the successful cell growth along with PDO production was achieved with the maximum PDO concentration of 1.5 mM by 15 days cultivation. The productivity of 1,3-PDO was approximately 0.1mM/day. The growth associated production of 1,3- PDO allows us to employ a continuous mode operation for 1,3-PDO production using ALPBS.

Therefore, the continuous production of 1,3-PDO was then carried out using the ALPB. The medium used was the BG-11 (pH 7.5 by adding HEPES). The operation was started by a batch culture for 11 days and then shifted to continuous operation for 60 days. Dilution rates were increased step by step. The ALPB was successfully operated in a continuous mode, showing 1.5 mM of 1,3-PDO concentration with the productivity of 0.1 mM/day.

In order to improve the 1,3-PDO productivity in the continuous process, pH-stat system using CO₂ for pH control was introduced into the ALPB. The pH-stat system was expected to increase 1,3-PDO production by the supplemental CO₂ supply as well as the optimal pH control for 1,3-PDO production. By introducing the pH-stat system, the pH in the ALPB was precisely controlled. The operation of ALPB with pH-stat system was stable and successfully operated for 60 days. The maximum productivity of 1,3-PDO in the continuous cultivation controlled at pH 8.0 was significantly increased to 0.23 mM/day and the pH-stat system was found to be effective to improve the 1,3-PDO production in the continuous operation.

Thus, the effective continuous production of 1,3-PDO using genetically engineered S. elongatus was achieved by employing the ALPB with pH-stat system.

Reference

(1) Hirokawa, Y., et al., Cyanobacterial production of 1,3-propanediol directly from carbon dioxide using a synthetic metabolic pathway Metab. Eng. 34, 97-103 (2016)

(2) Hirokawa, Y., et al., Metabolic engineering of Synechococcus elongatus PCC 7942 for improvement of 1,3-propanediol and glycerol production based on in silico simulation of metabolic flux distribution Microbial Cell Factories 16, 212 (2017)