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Transcriptional analysis of Amorphotheca resinae ZN1 on biological degradation of furfural and 5-hydroxymethylfurfural derived from lignocellulose pretreatment.

Wang X, Gao Q, Bao J - Biotechnol Biofuels (2015)

Bottom Line: During the detoxification process, A. resinae ZN1 firstly reduced furfural or HMF into furfuryl alcohol or HMF alcohol, and then oxidized into furoic acid or HMF acid through furan aldehyde as the intermediate at low concentration level.Two Zn-dependent alcohol dehydrogenase genes and five AKR/ARI genes were found to be responsible for the furfural and HMF conversion to their corresponding alcohols.The genes responsible for the furfural and HMF degradation to the corresponding alcohols and acids in A. resinae ZN1 were identified based on the analysis of the genome annotation, the gene transcription data and the inhibitor conversion results.

View Article: PubMed Central - PubMed

Affiliation: State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, 130 Meilong Road, Shanghai, 200237 China.

ABSTRACT

Background: Furfural and 5-hydroxymethylfurfural (HMF) are the two major inhibitor compounds generated from lignocellulose pretreatment, especially for dilute acid, steam explosion, neutral hot water pretreatment methods. The two inhibitors severely inhibit the cell growth and metabolism of fermenting strains in the consequent bioconversion step. The biodetoxification strain Amorphotheca resinae ZN1 has demonstrated its extraordinary capacity of fast and complete degradation of furfural and HMF into corresponding alcohol and acid forms. The elucidation of degradation metabolism of A. resinae ZN1 at molecular level will facilitate the detoxification of the pretreated lignocellulose biomass and provide the metabolic pathway information for more powerful biodetoxification strain development.

Results: Amorphotheca resinae ZN1 was able to use furfural or HMF as the sole carbon source for cell growth. During the detoxification process, A. resinae ZN1 firstly reduced furfural or HMF into furfuryl alcohol or HMF alcohol, and then oxidized into furoic acid or HMF acid through furan aldehyde as the intermediate at low concentration level. The cell mass measurement suggested that furfural was more toxic to A. resinae ZN1 than HMF. In order to identify the degradation mechanism of A. resinae ZN1, transcription levels of 137 putative genes involved in the degradation of furfural and HMF in A. resinae ZN1 were investigated using the real-time quantitative PCR (qRT-PCR) method under the stress of furfural and HMF, as well as the stress of their secondary metabolites, furfuryl alcohol and HMF alcohol. Two Zn-dependent alcohol dehydrogenase genes and five AKR/ARI genes were found to be responsible for the furfural and HMF conversion to their corresponding alcohols. For the conversion of the two furan alcohols to the corresponding acids, three propanol-preferring alcohol dehydrogenase genes, one NAD(P)(+)-depending aldehyde dehydrogenase gene, or two oxidase genes with free oxygen as the substrate were identified under aerobic condition.

Conclusions: The genes responsible for the furfural and HMF degradation to the corresponding alcohols and acids in A. resinae ZN1 were identified based on the analysis of the genome annotation, the gene transcription data and the inhibitor conversion results. These genetic resources provided the important information for understanding the mechanism of furfural and HMF degradation and modification of high tolerant strains used for biorefinery processing.

No MeSH data available.


Related in: MedlinePlus

Comparison of transcription levels for selected relevant genes in A. resinae ZN1 in response to 1 g/L of furfuryl alcohol or HMF alcohol. Quantitative expression level for each gene is log2 transformed from raw fold changes against that at 0 h. Red indicates up-regulated expression and blue for down-regulated expression as indicated by a color bar at the figure right. Expression data marked with green indicate significantly up-regulated (more than 200-fold) data. The different relevant genes are listed on the figure left and the color from top to bottom indicates different categories of genes: purpleADH genes, blueAKR/ARI genes, orangeALDH genes, green oxidase genes
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Fig5: Comparison of transcription levels for selected relevant genes in A. resinae ZN1 in response to 1 g/L of furfuryl alcohol or HMF alcohol. Quantitative expression level for each gene is log2 transformed from raw fold changes against that at 0 h. Red indicates up-regulated expression and blue for down-regulated expression as indicated by a color bar at the figure right. Expression data marked with green indicate significantly up-regulated (more than 200-fold) data. The different relevant genes are listed on the figure left and the color from top to bottom indicates different categories of genes: purpleADH genes, blueAKR/ARI genes, orangeALDH genes, green oxidase genes

Mentions: Amorphotheca resinae ZN1 degrades furfural or HMF into furfuryl alcohol or HMF alcohol, and further into furoic acid or HMF acid under aerobic condition. Furoic acid and HMF acid are commonly considered as almost non-toxic thus the biodetoxification ends with the furan acids formation [26, 30]. However, the degradation mechanism of furan alcohols to furan aldehydes and acids has not been clarified, although furfuryl alcohol and HMF alcohol are less toxic than furan aldehydes and still considered as inhibitors on the cell growth and metabolism of fermenting strains. In this study, the transcription response of A. resinae ZN1 to furfuryl alcohol and HMF alcohol were quantified using qRT-PCR. The RNA samples were collected after cultured for 48 h under the stress of furfuryl alcohol or HMF alcohol, respectively, at the time point the degradation of the alcohols was clearly observed. The transcription levels of 80 ADH genes, 21 AKR/ARI genes on the oxidation pathways of furan alcohols to furan aldehydes, as well as 20 ALDH genes, 16 oxidase genes involved in the conversion of furan aldehydes to furan acids, were examined as shown in Fig. 5. Under the stress of furfuryl alcohol, 29 ADH genes, 5 AKR/ARI genes, 5 ALDH genes and 7 oxidase genes were up-regulated by more than twofold, with only 3 ADH gene were down-regulated with the fold change less than 0.5. Under the stress of HMF alcohol, 30 ADH genes, 6 AKR/ARI genes, 8 ALDH genes and 8 oxidase genes were up-regulated, with 6 ADH genes, 1 AKR/ARI genes, and 1 ALDH genes were down-regulated. The total number of the up-regulated genes under furfuryl alcohol stress was close to that under HMF alcohol stress (46:52).Fig. 5


Transcriptional analysis of Amorphotheca resinae ZN1 on biological degradation of furfural and 5-hydroxymethylfurfural derived from lignocellulose pretreatment.

Wang X, Gao Q, Bao J - Biotechnol Biofuels (2015)

Comparison of transcription levels for selected relevant genes in A. resinae ZN1 in response to 1 g/L of furfuryl alcohol or HMF alcohol. Quantitative expression level for each gene is log2 transformed from raw fold changes against that at 0 h. Red indicates up-regulated expression and blue for down-regulated expression as indicated by a color bar at the figure right. Expression data marked with green indicate significantly up-regulated (more than 200-fold) data. The different relevant genes are listed on the figure left and the color from top to bottom indicates different categories of genes: purpleADH genes, blueAKR/ARI genes, orangeALDH genes, green oxidase genes
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4559888&req=5

Fig5: Comparison of transcription levels for selected relevant genes in A. resinae ZN1 in response to 1 g/L of furfuryl alcohol or HMF alcohol. Quantitative expression level for each gene is log2 transformed from raw fold changes against that at 0 h. Red indicates up-regulated expression and blue for down-regulated expression as indicated by a color bar at the figure right. Expression data marked with green indicate significantly up-regulated (more than 200-fold) data. The different relevant genes are listed on the figure left and the color from top to bottom indicates different categories of genes: purpleADH genes, blueAKR/ARI genes, orangeALDH genes, green oxidase genes
Mentions: Amorphotheca resinae ZN1 degrades furfural or HMF into furfuryl alcohol or HMF alcohol, and further into furoic acid or HMF acid under aerobic condition. Furoic acid and HMF acid are commonly considered as almost non-toxic thus the biodetoxification ends with the furan acids formation [26, 30]. However, the degradation mechanism of furan alcohols to furan aldehydes and acids has not been clarified, although furfuryl alcohol and HMF alcohol are less toxic than furan aldehydes and still considered as inhibitors on the cell growth and metabolism of fermenting strains. In this study, the transcription response of A. resinae ZN1 to furfuryl alcohol and HMF alcohol were quantified using qRT-PCR. The RNA samples were collected after cultured for 48 h under the stress of furfuryl alcohol or HMF alcohol, respectively, at the time point the degradation of the alcohols was clearly observed. The transcription levels of 80 ADH genes, 21 AKR/ARI genes on the oxidation pathways of furan alcohols to furan aldehydes, as well as 20 ALDH genes, 16 oxidase genes involved in the conversion of furan aldehydes to furan acids, were examined as shown in Fig. 5. Under the stress of furfuryl alcohol, 29 ADH genes, 5 AKR/ARI genes, 5 ALDH genes and 7 oxidase genes were up-regulated by more than twofold, with only 3 ADH gene were down-regulated with the fold change less than 0.5. Under the stress of HMF alcohol, 30 ADH genes, 6 AKR/ARI genes, 8 ALDH genes and 8 oxidase genes were up-regulated, with 6 ADH genes, 1 AKR/ARI genes, and 1 ALDH genes were down-regulated. The total number of the up-regulated genes under furfuryl alcohol stress was close to that under HMF alcohol stress (46:52).Fig. 5

Bottom Line: During the detoxification process, A. resinae ZN1 firstly reduced furfural or HMF into furfuryl alcohol or HMF alcohol, and then oxidized into furoic acid or HMF acid through furan aldehyde as the intermediate at low concentration level.Two Zn-dependent alcohol dehydrogenase genes and five AKR/ARI genes were found to be responsible for the furfural and HMF conversion to their corresponding alcohols.The genes responsible for the furfural and HMF degradation to the corresponding alcohols and acids in A. resinae ZN1 were identified based on the analysis of the genome annotation, the gene transcription data and the inhibitor conversion results.

View Article: PubMed Central - PubMed

Affiliation: State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, 130 Meilong Road, Shanghai, 200237 China.

ABSTRACT

Background: Furfural and 5-hydroxymethylfurfural (HMF) are the two major inhibitor compounds generated from lignocellulose pretreatment, especially for dilute acid, steam explosion, neutral hot water pretreatment methods. The two inhibitors severely inhibit the cell growth and metabolism of fermenting strains in the consequent bioconversion step. The biodetoxification strain Amorphotheca resinae ZN1 has demonstrated its extraordinary capacity of fast and complete degradation of furfural and HMF into corresponding alcohol and acid forms. The elucidation of degradation metabolism of A. resinae ZN1 at molecular level will facilitate the detoxification of the pretreated lignocellulose biomass and provide the metabolic pathway information for more powerful biodetoxification strain development.

Results: Amorphotheca resinae ZN1 was able to use furfural or HMF as the sole carbon source for cell growth. During the detoxification process, A. resinae ZN1 firstly reduced furfural or HMF into furfuryl alcohol or HMF alcohol, and then oxidized into furoic acid or HMF acid through furan aldehyde as the intermediate at low concentration level. The cell mass measurement suggested that furfural was more toxic to A. resinae ZN1 than HMF. In order to identify the degradation mechanism of A. resinae ZN1, transcription levels of 137 putative genes involved in the degradation of furfural and HMF in A. resinae ZN1 were investigated using the real-time quantitative PCR (qRT-PCR) method under the stress of furfural and HMF, as well as the stress of their secondary metabolites, furfuryl alcohol and HMF alcohol. Two Zn-dependent alcohol dehydrogenase genes and five AKR/ARI genes were found to be responsible for the furfural and HMF conversion to their corresponding alcohols. For the conversion of the two furan alcohols to the corresponding acids, three propanol-preferring alcohol dehydrogenase genes, one NAD(P)(+)-depending aldehyde dehydrogenase gene, or two oxidase genes with free oxygen as the substrate were identified under aerobic condition.

Conclusions: The genes responsible for the furfural and HMF degradation to the corresponding alcohols and acids in A. resinae ZN1 were identified based on the analysis of the genome annotation, the gene transcription data and the inhibitor conversion results. These genetic resources provided the important information for understanding the mechanism of furfural and HMF degradation and modification of high tolerant strains used for biorefinery processing.

No MeSH data available.


Related in: MedlinePlus