A seawater-based open and continuous process for polyhydroxyalkanoates production by recombinant Halomonas campaniensis LS21 grown in mixed substrates

Abstract

Background: High-cost production of bioplastics polyhydroxyalkanoates (PHA) is a major concern for their large scale application. In order to produce PHA economically, new technology must be developed to reduce costs on energy consumption, fresh water and substrate usages. It is also important to conduct the PHA production process in a continuous way rather than in a batch process. Results: A halophile Halomonas campaniensis strain LS21 was isolated to allow the development of a sea water based open and continuous process for PHA production utilizing mixed substrates consisting of mostly cellulose, starch, lipids and proteins. To study the feasibilities of open and long-term cultivation as well as genetic manipulation of this strain, polyhydroxybutyrate (PHB), the first member of the diverse PHA family, was taken as an example for the application of H. campaniensis LS21 in a robust and long lasting fermentation process. Wild type and recombinant H. campaniensis LS21 containing a PHB synthesis genes phbCAB were allowed respectively to grow in artificial seawater containing mixed substrates similar to kitchen wastes, including soluble and insoluble cellulose, proteins, fats, fatty acids and starch for 65 days without interruption. In the presence of 27 g/L NaCl under a pH around 10 at 37°C, the recombinant produced approximately 70% PHB and the wild type 26% during the 65 days fermentation process without infection. H. campaniensis LS21 secreted extracellular amylase, lipase, protease and cellulase simultaneously during the whole process to allow consumption of the mixed substrates. The recombinant was also found to stably maintain the phbCAB plasmid over the entire 65 days process. Conclusions:The seawater based open and continuous process based on halophilic Halomonas campaniensis LS21 allowed the applications of kitchen wastes like mixed substrates as nutrients for production of bioplastic PHB. This study demonstrates the advantages of this technology in terms of energy saving (non-sterilization), seawater based (not fresh water needed), long-lasting and continuous open processing (against batch process), and low cost substrates (non-food mixed substrates). Combined with its ease of genetic manipulation, Halomonas campaniensis LS21 could be developed into a platform for low cost production of chemicals, materials and biofuels.

Production of Polyhydroxyalkanoate (PHA) Biopolyesters by Extremophiles?

“…Until the end of the cultivation, the plasmid carrying phbCAB genes maintained stable in the recombinant cells. Combined with its expedient susceptibility for genetic improvement, H. campaniensis LS21 definitely might constitute a powerful cellular factory for cost-and energy-efficient production of PHA or extremozymes from inexpensive feedstocks [72].…”

Genome Structure ofBacillus cereustsu1 and Genes Involved in Cellulose Degradation and Poly-3-Hydroxybutyrate Synthesis

“…The high cost associated with the production of PHAs remains a major barrier for large scale use of bioplastics [58]. In order to produce PHAs economically, a reliable and economical supply of raw material is essential; meanwhile, new bacterial strains and genes need to be discovered in order (Invitrogen, CA); recombinant proteins were expressed in DE3 cells.…”

A novel cell autolysis system for cost-competitive downstream processing

“…E. coli cells were grown in LB10 with the addition of 25 mg/L chloramphenicol (Sigma-Aldrich, Germany) from a 25 mg/mL stock solution in ethanol to promote plasmid retention and incubated at 37°C under constant orbital shaking at 200 rpm. The Halomonas LS21 strains were derived from the wild-type strain published earlier (Yue et al 2014), deposited in the China General Microbiological Culture Collection Center under the code CGMCC No. 6593 and were grown in LB40 media, but otherwise the same as described for E. coli.…”