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Biotechnol Biofuels. 2019 Feb 27;12:41. doi: 10.1186/s13068-019-1381-3. eCollection 2019.

Engineering Corynebacterium glutamicum to produce the biogasoline isopentenol from plant biomass hydrolysates.

Author information

1
1Graduate School of Advanced Integrated Studies in Human Survivability, Kyoto University, Sakyo-ku, Kyoto, Japan.
2
2Japan Society for the Promotion of Science, Sakyo-ku, Kyoto, Japan.
3
3Joint BioEnergy Institute, Emeryville, CA USA.
4
5Biological Systems and Engineering Division, Lawrence Berkeley National Laboratory, Berkeley, CA USA.
5
4Biomass Science and Conversion Technology Department, Sandia National Laboratories, 7011 East Avenue, Livermore, CA 94550 USA.
6
6Environmental Genomics and Systems Biology Division, Lawrence Berkeley National Laboratory, Berkeley, CA USA.
#
Contributed equally

Abstract

Background:

Many microbes used for the rapid discovery and development of metabolic pathways have sensitivities to final products and process reagents. Isopentenol (3-methyl-3-buten-1-ol), a biogasoline candidate, has an established heterologous gene pathway but is toxic to several microbial hosts. Reagents used in the pretreatment of plant biomass, such as ionic liquids, also inhibit growth of many host strains. We explored the use of Corynebacterium glutamicum as an alternative host to address these constraints.

Results:

We found C. glutamicum ATCC 13032 to be tolerant to both the final product, isopentenol, as well to three classes of ionic liquids. A heterologous mevalonate-based isopentenol pathway was engineered in C. glutamicum. Targeted proteomics for the heterologous pathway proteins indicated that the 3-hydroxy-3-methylglutaryl-coenzyme A reductase protein, HmgR, is a potential rate-limiting enzyme in this synthetic pathway. Isopentenol titers were improved from undetectable to 1.25 g/L by combining three approaches: media optimization; substitution of an NADH-dependent HmgR homolog from Silicibacter pomeroyi; and development of a C. glutamicum ∆poxB ∆ldhA host chassis.

Conclusions:

We describe the successful expression of a heterologous mevalonate-based pathway in the Gram-positive industrial microorganism, C. glutamicum, for the production of the biogasoline candidate, isopentenol. We identified critical genetic factors to harness the isopentenol pathway in C. glutamicum. Further media and cultivation optimization enabled isopentenol production from sorghum biomass hydrolysates.

KEYWORDS:

3-Methyl-3-buten-1-ol; Corynebacterium glutamicum; HmgR; Hydrolysate; Ionic liquid pretreatment; Isopentenol; Isoprenol; Sorghum

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