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Metabolic engineering of Acinetobacter baylyi ADP1 for removal of Clostridium butyricum growth inhibitors produced from lignocellulosic hydrolysates.

Kannisto MS, Mangayil RK, Shrivastava-Bhattacharya A, Pletschke BI, Karp MT, Santala VP - Biotechnol Biofuels (2015)

Bottom Line: Pretreatment of lignocellulosic biomass can produce inhibitory compounds that are harmful for microorganisms used in the production of biofuels and other chemicals from lignocellulosic sugars.Formate was consumed during growth on acetate and by stationary phase cells, and this was enhanced in the presence of a common aromatic inhibitor of lignocellulosic hydrolysates, 4-hydroxybenzoate.Because of these encouraging results, we believe that A. baylyi ADP1 is a promising candidate for the detoxification of lignocellulosic hydrolysates for bioprocesses.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry and Bioengineering, Tampere University of Technology, Korkeakoulunkatu 8, Tampere, Finland.

ABSTRACT

Background: Pretreatment of lignocellulosic biomass can produce inhibitory compounds that are harmful for microorganisms used in the production of biofuels and other chemicals from lignocellulosic sugars. Selective inhibitor removal can be achieved with biodetoxification where microorganisms catabolize the inhibitors without consuming the sugars. We engineered the strictly aerobic Acinetobacter baylyi ADP1 for detoxification of lignocellulosic hydrolysates by removing the gene for glucose dehydrogenase, gcd, which catalyzes the first step in its glucose catabolism.

Results: The engineered A. baylyi ADP1 strain was shown to be incapable of consuming the main sugar components of lignocellulosic hydrolysates, i.e., glucose, xylose, and arabinose, but rapidly utilized acetate and formate. Formate was consumed during growth on acetate and by stationary phase cells, and this was enhanced in the presence of a common aromatic inhibitor of lignocellulosic hydrolysates, 4-hydroxybenzoate. The engineered strain tolerated glucose well up to 70 g/l, and the consumption of glucose, xylose, or arabinose was not observed in prolonged cultivations. The engineered strain was applied in removal of oxygen, a gaseous inhibitor of anaerobic fermentations. Co-cultivation with the A. baylyi ADP1 gcd knockout strain under initially aerobic conditions allowed the strictly anaerobic Clostridium butyricum to grow and produce hydrogen (H2) from sugars of the enzymatic rice straw hydrolysate.

Conclusions: We demonstrated that the model organism of bacterial genetics and metabolism, A. baylyi ADP1, could be engineered to be an efficient biodetoxification strain of lignocellulosic hydrolysates. Only one gene knockout was required to completely eliminate sugar consumption and the strain could be used in production of anaerobic conditions for the strictly anaerobic hydrogen producer, C. butyricum. Because of these encouraging results, we believe that A. baylyi ADP1 is a promising candidate for the detoxification of lignocellulosic hydrolysates for bioprocesses.

No MeSH data available.


Related in: MedlinePlus

Growth and consumption of d-glucose, d-xylose, l-arabinose, acetate, formate, and levulinate of wild-type A. baylyi ADP1 (a) and gcd knockout strain (b). Cultivations were carried out in triplicate and the results are shown as averages with error bars representing standard deviations
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Fig1: Growth and consumption of d-glucose, d-xylose, l-arabinose, acetate, formate, and levulinate of wild-type A. baylyi ADP1 (a) and gcd knockout strain (b). Cultivations were carried out in triplicate and the results are shown as averages with error bars representing standard deviations

Mentions: In order to characterize the growth of A. baylyi ADP1 and gcd knockout strain on a mixture of sugars and organic acids commonly found in most lignocellulosic hydrolysates, we cultivated the cells on approximately 10 mM of glucose, xylose, arabinose, formate, acetate, and levulinate (Fig. 1). The wild-type cells consumed glucose, formate, and acetate simultaneously. Acetate was consumed the most rapidly, and the growth rate was the highest when it was available as a carbon source, but it did not seem to repress catabolism of glucose or formate to great extent. Xylose and arabinose were oxidized less preferably than glucose. The gcd knockout strain grew slightly faster (µ = 0.58 ± 0.01 h−1) than the wild-type strain (µ = 0.50 ± 0.02 h−1) during the beginning (2–6 h) of the cultivation, without consuming any of the sugars, but its growth ceased when acetate was depleted. However, formate consumption continued after this point and it seems that it cannot be used as a sole carbon source for growth but can be still be consumed by stationary phase cells. Acetate consumption was slightly faster than with wild-type cells but formate was consumed less rapidly. Neither strain was able to catabolize levulinate and this molecule was therefore excluded from subsequent experiments.Fig. 1


Metabolic engineering of Acinetobacter baylyi ADP1 for removal of Clostridium butyricum growth inhibitors produced from lignocellulosic hydrolysates.

Kannisto MS, Mangayil RK, Shrivastava-Bhattacharya A, Pletschke BI, Karp MT, Santala VP - Biotechnol Biofuels (2015)

Growth and consumption of d-glucose, d-xylose, l-arabinose, acetate, formate, and levulinate of wild-type A. baylyi ADP1 (a) and gcd knockout strain (b). Cultivations were carried out in triplicate and the results are shown as averages with error bars representing standard deviations
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4666034&req=5

Fig1: Growth and consumption of d-glucose, d-xylose, l-arabinose, acetate, formate, and levulinate of wild-type A. baylyi ADP1 (a) and gcd knockout strain (b). Cultivations were carried out in triplicate and the results are shown as averages with error bars representing standard deviations
Mentions: In order to characterize the growth of A. baylyi ADP1 and gcd knockout strain on a mixture of sugars and organic acids commonly found in most lignocellulosic hydrolysates, we cultivated the cells on approximately 10 mM of glucose, xylose, arabinose, formate, acetate, and levulinate (Fig. 1). The wild-type cells consumed glucose, formate, and acetate simultaneously. Acetate was consumed the most rapidly, and the growth rate was the highest when it was available as a carbon source, but it did not seem to repress catabolism of glucose or formate to great extent. Xylose and arabinose were oxidized less preferably than glucose. The gcd knockout strain grew slightly faster (µ = 0.58 ± 0.01 h−1) than the wild-type strain (µ = 0.50 ± 0.02 h−1) during the beginning (2–6 h) of the cultivation, without consuming any of the sugars, but its growth ceased when acetate was depleted. However, formate consumption continued after this point and it seems that it cannot be used as a sole carbon source for growth but can be still be consumed by stationary phase cells. Acetate consumption was slightly faster than with wild-type cells but formate was consumed less rapidly. Neither strain was able to catabolize levulinate and this molecule was therefore excluded from subsequent experiments.Fig. 1

Bottom Line: Pretreatment of lignocellulosic biomass can produce inhibitory compounds that are harmful for microorganisms used in the production of biofuels and other chemicals from lignocellulosic sugars.Formate was consumed during growth on acetate and by stationary phase cells, and this was enhanced in the presence of a common aromatic inhibitor of lignocellulosic hydrolysates, 4-hydroxybenzoate.Because of these encouraging results, we believe that A. baylyi ADP1 is a promising candidate for the detoxification of lignocellulosic hydrolysates for bioprocesses.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry and Bioengineering, Tampere University of Technology, Korkeakoulunkatu 8, Tampere, Finland.

ABSTRACT

Background: Pretreatment of lignocellulosic biomass can produce inhibitory compounds that are harmful for microorganisms used in the production of biofuels and other chemicals from lignocellulosic sugars. Selective inhibitor removal can be achieved with biodetoxification where microorganisms catabolize the inhibitors without consuming the sugars. We engineered the strictly aerobic Acinetobacter baylyi ADP1 for detoxification of lignocellulosic hydrolysates by removing the gene for glucose dehydrogenase, gcd, which catalyzes the first step in its glucose catabolism.

Results: The engineered A. baylyi ADP1 strain was shown to be incapable of consuming the main sugar components of lignocellulosic hydrolysates, i.e., glucose, xylose, and arabinose, but rapidly utilized acetate and formate. Formate was consumed during growth on acetate and by stationary phase cells, and this was enhanced in the presence of a common aromatic inhibitor of lignocellulosic hydrolysates, 4-hydroxybenzoate. The engineered strain tolerated glucose well up to 70 g/l, and the consumption of glucose, xylose, or arabinose was not observed in prolonged cultivations. The engineered strain was applied in removal of oxygen, a gaseous inhibitor of anaerobic fermentations. Co-cultivation with the A. baylyi ADP1 gcd knockout strain under initially aerobic conditions allowed the strictly anaerobic Clostridium butyricum to grow and produce hydrogen (H2) from sugars of the enzymatic rice straw hydrolysate.

Conclusions: We demonstrated that the model organism of bacterial genetics and metabolism, A. baylyi ADP1, could be engineered to be an efficient biodetoxification strain of lignocellulosic hydrolysates. Only one gene knockout was required to completely eliminate sugar consumption and the strain could be used in production of anaerobic conditions for the strictly anaerobic hydrogen producer, C. butyricum. Because of these encouraging results, we believe that A. baylyi ADP1 is a promising candidate for the detoxification of lignocellulosic hydrolysates for bioprocesses.

No MeSH data available.


Related in: MedlinePlus