Abstract
Most whole cell biocatalysts have some problems with yields and productivities because of various metabolites produced as byproducts and limitations of substrate uptake. We propose a psychrophile-based simple biocatalyst for efficient bio-production using mesophilic enzymes expressed in psychrophilic Shewanella livingstonensis Ac10 cells whose basic metabolism was inactivated by heat treatment. The 45°C heat-treated cells expressing lacZ showed maximum beta-galactosidase activity as well as chloroform/SDS-treated cells to increase membrane permeability. The fluorescent dye 5-cyano-2,3-ditolyl-tetrazolium chloride staining indicated that most basic metabolism of Ac10 was lost by heat treatment at 45˚C for 10 min. The simple biocatalyst was applied for 3-HPA production by using Klebsiella pneumoniae dhaB genes. 3-HPA was stoichiometrically produced with the complete consumption of glycerol at a high production rate of 8.85 mmol 3-HPA/g dry cell/h. The amount of 3-HPA production increased by increasing the concentrations of biocatalyst and glycerol. Furthermore, it could convert biodiesel-derived crude glycerol to 3-HPA.
Highlights
Microorganisms are used as platform cell factories to produce various building block chemicals from renewable resources
We evaluated the activity of recombinant E. coli β-galactosidase expressed in S. livingstonensis Ac10 and basic metabolism of Ac10 when cells were treated by heat, and applied it to the simple biocatalyst for 3-HPA production from glycerol by using
Heat treatment condition for simple biocatalyst To evaluate the effect of heat treatment on S. livingstonenesis Ac10, we measured the β-galactosidase activity of the recombinant Ac10 cells in which the E. coli lacZ gene was expressed under the control of a tac promoter
Summary
Microorganisms are used as platform cell factories to produce various building block chemicals from renewable resources. Thermostable enzymes expressed in Escherichia coli have achieved efficient bio-conversions with nearly 100% yields (Iwamoto et al 2007; Ye et al 2012). We considered the construction of a simple biocatalyst using psychrophilic bacterial cells (psychrophile-based simple biocatalyst) instead of E. coli cells. This biocatalyst has significant advantages in the use of abundant mesophilic enzymes for designing bio-conversion pathways, in addition to thermostable enzymes because the use of psychrophilic hosts allows the heat treatment temperature for inactivation of basic metabolisms to be lower than that of E. coli based simple biocatalysts. Psychrophile-based simple biocatalysts may be able to exploit most enzymes to produce chemicals. Decreasing the heat treatment temperature to an ordinary temperature limits concerns about the excess disruption of the cellular structure and thermal decomposition of coenzymes, such as NADP+ and NADPH, which are problems in E. coli-based simple biocatalysts (Ye et al 2013)
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