Engineering a methanol-tolerant methanotroph for conversion of methane and methanol to isoprenoids

#methane #energy #ethanol #alberta

Methane and methanol are inexpensive and sustainable carbon feedstocks that can be used by methanotrophic bacteria as sole sources of carbon and energy and can be converted into value-added products. On the other hand, isoprenoids are valuable compounds that can be used for high performance fuels, flavoring agents and pharmaceuticals or for precursors to such products; however, methanotrophs do not readily produce isoprenoids. Methylomicrobium album BG8 is a methanotroph with high methanol tolerance and has been grown to high density in fed-batch operation. In this work, M. album BG8 is being modified to convert methane and methanol into isoprenoids. M. album BG8 contains the doxp pathway, which produces isopentenyl diphosphate (IPP) and dimethylallyl diphosphate (DMAPP), two isoprenoid precursors, but it lacks enzymes which are required to transform these compounds into isoprenoids. In addition, works in other bacterial systems suggest low expression of the native doxp pathway leading to a potential bottleneck in the generation of isoprenoids. In this work, conjugation was used to modify M. album BG8 with plasmids for high and low expression levels of the genes involved in isoprenoid production. A library of constitutive and inducible promoters was developed using green fluorescent protein (GFP) and nanoluc as markers. Co-transformation of high and low expression vectors are being performed to improve the flux through the doxp pathway and, eventually, isoprenoid synthesis. This work provides a basis for the production of isoprenoids from methane and methanol by bacteria, an approach with great economic and environmental potential.