Metabolic Engineering of Acinetobacter baylyi ADP1 for Improved Growth on Gluconate and Glucose

Kannisto, Matti; Aho, Tommi; Karp, Matti; Santala, Ville

doi:10.1128/aem.01837-14

Cited by 23 publications

(13 citation statements)

References 32 publications

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“…More recently, an optimized alkane production pathway was integrated in the strain (Lehtinen, Virtanen, et al, ). Microbial wax esters and alkanes have previously been produced using carbon sources such as glucose and organic acids (Cao et al, ; Fatma et al, ; Kannisto, Aho, Karp, & Santala, ; Lehtinen, Efimova, et al, ; Lehtinen, Efimova, et al, ; Lehtinen, Santala, et al, ; Salmela, Lehtinen, Efimova, Santala, & Mangayil, ; Santala, Efimova, Karp, & Santala, ; Santala, Efimova, & Santala, ; Schirmer, Rude, Li, Popova, & del Cardayre, ).…”

Section: Introductionmentioning

confidence: 99%

Alkane and wax ester production from lignin‐related aromatic compounds

Salmela

Lehtinen

Efimova

et al. 2019

Biotech & Bioengineering

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Lignin has potential as a sustainable feedstock for microbial production of industrially relevant molecules. However, the required lignin depolymerization yields a heterogenic mixture of aromatic monomers that are challenging substrates for the microorganisms commonly used in the industry. Here, we investigated the properties of lignin-related aromatic compounds (LRAs), namely coumarate, ferulate, and caffeate, in the synthesis of biomass and products in an LRA-utilizing bacterial host Acinetobacter baylyi ADP1.The biosynthesis products, wax esters, and alkanes are relevant compounds for the chemical and fuel industries. Here, wax esters were produced by a native pathway of ADP1, whereas alkanes were produced by a synthetic pathway introduced to the host.Using individual LRAs as substrates, the growth and product formation were monitored with internal biosensors and off-line analytics. Of the tested LRAs, coumarate was the most propitious in terms of product synthesis. Wax esters were produced from coumarate with yield and titer of 37 mg/g coumarate and 202 mg/L, whereas alkanes were produced with a yield of 62.3 µg /g coumarate and titer of 152 µg/L. This study demonstrates the microbial preference for certain LRAs and highlights the potential of A. baylyi ADP1 as a host for LRA upgrading to value-added products. K E Y W O R D SAcinetobacter baylyi ADP1, alkane, lignin, wax ester

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Section: Introductionmentioning

confidence: 99%

Alkane and wax ester production from lignin‐related aromatic compounds

Salmela

Lehtinen

Efimova

et al. 2019

Biotech & Bioengineering

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“…It was previously demonstrated that a knock-out of the isocitrate lyase in Pseudomonas putida improved the production of PHAs when grown on gluconate, apparently for providing surplus acetyl-CoA for the PHA synthesis ( Klinke et al, 2000 ). Some bacteria also exhibit an alternative pathway for glycolysis, such as the modified Entner-Doudoroff pathway of certain Acinetobacter strains ( Young et al, 2005 , Kannisto et al, 2014 ). An interesting feature of the glycolysis of Acinetobacter is the oxidation of glucose to gluconate prior to the transport to cells.…”

Section: Introductionmentioning

confidence: 99%

Dynamic decoupling of biomass and wax ester biosynthesis in Acinetobacter baylyi by an autonomously regulated switch

Santala

Efimova

Santala

2018

Metabolic Engineering Communications

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For improving the microbial production of fuels and chemicals, gene knock-outs and overexpression are routinely applied to intensify the carbon flow from substrate to product. However, their possibilities in dynamic control of the flux between the biomass and product synthesis are limited, whereas dynamic metabolic switches can be used for optimizing the distribution of carbon and resources. The production of single cell oils is especially challenging, as the synthesis is strictly regulated, competes directly with biomass, and requires defined conditions, such as nitrogen limitation. Here, we engineered a metabolic switch for redirecting carbon flow from biomass to wax ester production in Acinetobacter baylyi ADP1 using acetate as a carbon source. Isocitrate lyase, an essential enzyme for growth on acetate, was expressed under an arabinose inducible promoter. The autonomous downregulation of the expression is based on the gradual oxidation of the arabinose inducer by a glucose dehydrogenase gcd. The depletion of the inducer, occurring simultaneously to acetate consumption, switches the cells from a biomass mode to a lipid synthesis mode, enabling the efficient channelling of carbon to wax esters in a simple batch culture. In the engineered strain, the yield and titer of wax esters were improved by 3.8 and 3.1 folds, respectively, over the control strain. In addition, the engineered strain accumulated wax esters 19% of cell dry weight, being the highest reported among microbes. The study provides important insights into the dynamic engineering of the biomass-dependent synthesis pathways for the improved production of biocompounds from low-cost and sustainable substrates.

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“…A. baylyi ADP1 is a promising chassis for genetic studies, metabolic engineering, and lipid production [26–30]. The strain naturally produces storage lipids, and thus the central metabolism is wired for the efficient production of fatty acyl-derived molecules.…”

Section: Introductionmentioning

confidence: 99%

Production of alkanes from CO2 by engineered bacteria

Lehtinen

Virtanen

Santala

et al. 2018

Preprint

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BackgroundMicrobial biosynthesis of alkanes is considered a promising method for the sustainable production of drop-in fuels and chemicals. Carbon dioxide would be an ideal carbon source for these production systems, but efficient production of long carbon chains from CO2 is difficult to achieve in a single organism. A potential solution is to employ acetogenic bacteria for the reduction of CO2 to acetate, and engineer a second organism to convert the acetate into long-chain hydrocarbons.ResultsIn this study, we demonstrate alkane production from CO2 by a system combining the acetogen Acetobacterium woodii and a non-native alkane producer Acinetobacter baylyi ADP1 engineered for alkane production. Nine synthetic two-step alkane biosynthesis pathways consisting of different aldehyde- and alkane-producing enzymes were combinatorically constructed and expressed in A. baylyi. The aldehyde-producing enzymes studied were AAR from Synechococcus elongatus, Acr1 from A. baylyi, and Ramo, a putative dehydrogenase, from Nevskia ramosa. The alkane-producing enzymes were ADOs from S. elongatus and Nostoc punctiforme, and CER1 from Arabidopsis thaliana. The performance of the pathways was evaluated with a twin-layer biosensor, which allowed the monitoring of both the intermediate, fatty aldehyde, as well as the alkane production. The highest alkane production, as indicated by the biosensor, was achieved with a pathway consisting of AAR and ADO from S. elongatus. The performance of this pathway was further improved by balancing the relative expression levels of the enzymes in order to limit the accumulation of the intermediate fatty aldehyde. Finally, the acetogen A. woodii was used to produce acetate from CO2 and H2, and the acetate was used for alkane production by the engineered A. baylyi, thereby leading to the net production of long-chain alkanes from CO2.ConclusionsA modular system for the production of drop-in liquid fuels from CO2 was demonstrated. Among the studied synthetic pathways, the combination of ADO and AAR from S. elongatus was found to be the most efficient in heterologous alkane production in A. baylyi. Furthermore, limiting the accumulation of the fatty aldehyde intermediate was found to be beneficial for the alkane production.

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Metabolic Engineering of Acinetobacter baylyi ADP1 for Improved Growth on Gluconate and Glucose

Cited by 23 publications

References 32 publications

Alkane and wax ester production from lignin‐related aromatic compounds

Alkane and wax ester production from lignin‐related aromatic compounds

Dynamic decoupling of biomass and wax ester biosynthesis in Acinetobacter baylyi by an autonomously regulated switch

Production of alkanes from CO2 by engineered bacteria

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