Novel microorganism and its use in lignocellulose detoxification
OwnerFrank Wouter KOOPMAN, Harald Johan Ruijssenaars +2inventors
InventorsFrank Wouter KOOPMAN, Harald Johan Ruijssenaars, Nick Johannes Petrus Wierckx, Johannes Hendrik De Winde
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Abstract
An isolated microorganism is provided which is Cupriavidus basilensis strain HMF14 Deposit number DSM 22875, and its use in a process for the in-situ detoxification of lignocelluloses hydrolysate.
Claims565
An isolated microorganism which is Cupriavidus basilensis strain HMF14 Deposit number DSM 22875.
A polypeptide having aldehyde dehydrogenase activity which comprises the amino acid sequence set out in SEQ ID NO:
- 15 or an amino acid sequence encoded by the nucleotide sequence of SEQ ID NO:
- 16 or a variant polypeptide thereof, wherein the variant has at least 64% sequence identity or more with the sequence set out in SEQ ID NO:
- 16.
A polynucleotide which comprises:
- (a) the nucleotide sequence set out in SEQ ID NO:
- 16;
- (b) a nucleotide sequence which hybridizes selectively with a polynucleotide being the reverse complement of SEQ ID NO:
- 16;
- (c) a nucleotide sequence having at least 64% sequence identity or more with the nucleotide sequence of SEQ ID NO:
- 16;
- (d) a fragment of a nucleotide sequence as defined in (a), (b) or (c) which is at least about 100 nucleotides in length;
- (e) a sequence which is degenerate as a result of the genetic code to a sequence as defined in any one of (a), (b), (c) or (d);
- (f) a nucleotide sequence which is the reverse complement of a nucleotide sequence as defined in (a), (b), (c), (d) or (e).
A polypeptide having LysR family transcriptional regulator activity which comprises the amino acid sequence set out in SEQ ID NO:
- 17 or an amino acid sequence encoded by the nucleotide sequence of SEQ ID NO:
- 18 or a variant polypeptide thereof, wherein the variant has at least 47% sequence identity or more with the sequence set out in SEQ ID NO:
- 17.
A polynucleotide which comprises:
- (a) the nucleotide sequence set out in SEQ ID NO:
- 18;
- (b) a nucleotide sequence which hybridizes selectively with a polynucleotide being the reverse complement of SEQ ID NO:
- 18;
- (c) a nucleotide sequence having at least 72% sequence identity or more with the nucleotide sequence of SEQ ID NO:
- 18;
- (d) a fragment of a nucleotide sequence as defined in (a), (b) or (c) which is at least about 100 nucleotides in length;
- (e) a sequence which is degenerate as a result of the genetic code to a sequence as defined in any one of (a), (b), (c) or (d);
- (f) a nucleotide sequence which is the reverse complement of a nucleotide sequence as defined in (a), (b), (c), (d) or (e).
A polypeptide having 2,5-furan-dicarboxylic acid decarboxylase 1 activity which comprises the amino acid sequence set out in SEQ ID NO:
- 19 or an amino acid sequence encoded by the nucleotide sequence of SEQ ID NO:
- 20 or a variant polypeptide thereof, wherein the variant has at least 54% sequence identity or more with the sequence set out in SEQ ID NO:
- 19.
A polynucleotide which comprises:
- (a) the nucleotide sequence set out in SEQ ID NO:
- 20;
- (b) a nucleotide sequence which hybridizes selectively with a polynucleotide being the reverse complement of SEQ ID NO:
- 20;
- (c) a nucleotide sequence having at least 66% sequence identity or more with the nucleotide sequence of SEQ ID NO:
- 20;
- (d) a fragment of a nucleotide sequence as defined in (a), (b) or (c) which is at least about 100 nucleotides in length;
- (e) a sequence which is degenerate as a result of the genetic code to a sequence as defined in any one of (a), (b), (c) or (d);
- (f) a nucleotide sequence which is the reverse complement of a nucleotide sequence as defined in (a), (b), (c), (d) or (e).
A polypeptide having 2,5-furan-dicarboxylic acid decarboxylase 2 activity which comprises the amino acid sequence set out in SEQ ID NO:
- 21 or an amino acid sequence encoded by the nucleotide sequence of SEQ ID NO:
- 22 or a variant polypeptide thereof, wherein the variant has at least 52% sequence identity or more with the sequence set out in SEQ ID NO:
- 21.
A polynucleotide which comprises:
- (a) the nucleotide sequence set out in SEQ ID NO:
- 22;
- (b) a nucleotide sequence which hybridizes selectively with a polynucleotide being the reverse complement of SEQ ID NO:
- 22;
- (c) a nucleotide sequence having at least 67% sequence identity or more with the nucleotide sequence of SEQ ID NO:
- 22;
- (d) a fragment of a nucleotide sequence as defined in (a), (b) or (c) which is at least about 100 nucleotides in length;
- (e) a sequence which is degenerate as a result of the genetic code to a sequence as defined in any one of (a), (b), (c) or (d);
- (f) a nucleotide sequence which is the reverse complement of a nucleotide sequence as defined in (a), (b), (c), (d) or (e).
A polypeptide having HMF/furfural oxidoreductase activity which comprises the amino acid sequence set out in SEQ ID NO:
- 25 or an amino acid sequence encoded by the nucleotide sequence of SEQ ID NO:
- 26 or a variant polypeptide thereof, wherein the variant has at least 45% sequence identity or more with the sequence set out in SEQ ID NO:
- 25.
A polynucleotide which comprises:
- (a) the nucleotide sequence set out in SEQ ID NO:
- 26;
- (b) a nucleotide sequence which hybridizes selectively with a polynucleotide being the reverse complement of SEQ ID NO:
- 26;
- (c) a nucleotide sequence having at least 66% sequence identity or more with the nucleotide sequence of SEQ ID NO:
- 26;
- (d) a fragment of a nucleotide sequence as defined in (a), (b) or (c) which is at least about 100 nucleotides in length;
- (e) a sequence which is degenerate as a result of the genetic code to a sequence as defined in any one of (a), (b), (c) or (d);
- (f) a nucleotide sequence which is the reverse complement of a nucleotide sequence as defined in (a), (b), (c), (d) or (e).
A polypeptide having LysR type transcriptional regulator activity which comprises the amino acid sequence set out in SEQ ID NO:
- 33 or an amino acid sequence encoded by the nucleotide sequence of SEQ ID NO:
- 34 or a variant polypeptide thereof, wherein the variant has at least 46% sequence identity or more with the sequence set out in SEQ ID NO:
- 33.
A polynucleotide which comprises:
- (a) the nucleotide sequence set out in SEQ ID NO:
- 34;
- (b) a nucleotide sequence which hybridizes selectively with a polynucleotide being the reverse complement of SEQ ID NO:
- 34;
- (c) a nucleotide sequence having at least 65% sequence identity or more with the nucleotide sequence of SEQ ID NO:
- 34;
- (d) a fragment of a nucleotide sequence as defined in (a), (b) or (c) which is at least about 100 nucleotides in length;
- (e) a sequence which is degenerate as a result of the genetic code to a sequence as defined in any one of (a), (b), (c) or (d);
- (f) a nucleotide sequence which is the reverse complement of a nucleotide sequence as defined in (a), (b), (c), (d) or (e).
A polypeptide having Furoyl-CoA dehydrogenase (large subunit) activity which comprises the amino acid sequence set out in SEQ ID NO:
- 35 or an amino acid sequence encoded by the nucleotide sequence of SEQ ID NO:
- 36 or a variant polypeptide thereof, wherein the variant has at least 54% sequence identity or more with the sequence set out in SEQ ID NO:
- 35.
A polynucleotide which comprises:
- (a) the nucleotide sequence set out in SEQ ID NO:
- 36;
- (b) a nucleotide sequence which hybridizes selectively with a polynucleotide being the reverse complement of SEQ ID NO:
- 36;
- (c) a nucleotide sequence having at least 66% sequence identity or more with the nucleotide sequence of SEQ ID NO:
- 36;
- (d) a fragment of a nucleotide sequence as defined in (a), (b) or (c) which is at least about 100 nucleotides in length;
- (e) a sequence which is degenerate as a result of the genetic code to a sequence as defined in any one of (a), (b), (c) or (d);
- (f) a nucleotide sequence which is the reverse complement of a nucleotide sequence as defined in (a), (b), (c), (d) or (e).
A polypeptide having Furoyl-CoA dehydrogenase (FAD binding subunit) activity which comprises the amino acid sequence set out in SEQ ID NO:
- 37 or an amino acid sequence encoded by the nucleotide sequence of SEQ ID NO:
- 38 or a variant polypeptide thereof, wherein the variant has at least 49% sequence identity or more with the sequence set out in SEQ ID NO:
- 37.
A polynucleotide which comprises:
- (a) the nucleotide sequence set out in SEQ ID NO:
- 38;
- (b) a nucleotide sequence which hybridizes selectively with a polynucleotide being the reverse complement of SEQ ID NO:
- 38;
- (c) a nucleotide sequence having at least 71% sequence identity or more with the nucleotide sequence of SEQ ID NO:
- 38;
- (d) a fragment of a nucleotide sequence as defined in (a), (b) or (c) which is at least about 100 nucleotides in length;
- (e) a sequence which is degenerate as a result of the genetic code to a sequence as defined in any one of (a), (b), (c) or (d);
- (f) a nucleotide sequence which is the reverse complement of a nucleotide sequence as defined in (a), (b), (c), (d) or (e).
A polypeptide having Furoyl-CoA dehydrogenase 2Fe-2S iron sulfur subunit activity which comprises the amino acid sequence set out in SEQ ID NO:
- 39 or an amino acid sequence encoded by the nucleotide sequence of SEQ ID NO:
- 40 or a variant polypeptide thereof, wherein the variant has at least 64% sequence identity or more with the sequence set out in SEQ ID NO:
- 39.
A polynucleotide which comprises:
- (a) the nucleotide sequence set out in SEQ ID NO:
- 40;
- (b) a nucleotide sequence which hybridizes selectively with a polynucleotide being the reverse complement of SEQ ID NO:
- 40;
- (c) a nucleotide sequence having at least 70% sequence identity or more with the nucleotide sequence of SEQ ID NO:
- 40;
- (d) a fragment of a nucleotide sequence as defined in (a), (b) or (c) which is at least about 100 nucleotides in length;
- (e) a sequence which is degenerate as a result of the genetic code to a sequence as defined in any one of (a), (b), (c) or (d);
- (f) a nucleotide sequence which is the reverse complement of a nucleotide sequence as defined in (a), (b), (c), (d) or (e).
A polypeptide having Furoyl-CoA synthetase activity which comprises the amino acid sequence set out in SEQ ID NO:
- 41 or an amino acid sequence encoded by the nucleotide sequence of SEQ ID NO:
- 42 or a variant polypeptide thereof, wherein the variant has at least 57% sequence identity or more with the sequence set out in SEQ ID NO:
- 41.
A polynucleotide which comprises:
- (a) the nucleotide sequence set out in SEQ ID NO:
- 42;
- (b) a nucleotide sequence which hybridizes selectively with a polynucleotide being the reverse complement of SEQ ID NO:
- 42;
- (c) a nucleotide sequence having at least 68% sequence identity or more with the nucleotide sequence of SEQ ID NO:
- 42;
- (d) a fragment of a nucleotide sequence as defined in (a), (b) or (c) which is at least about 100 nucleotides in length;
- (e) a sequence which is degenerate as a result of the genetic code to a sequence as defined in any one of (a), (b), (c) or (d);
- (f) a nucleotide sequence which is the reverse complement of a nucleotide sequence as defined in (a), (b), (c), (d) or (e).
A polypeptide having 2-oxoglutaroyl-CoA hydrolase activity which comprises the amino acid sequence set out in SEQ ID NO:
- 43 or an amino acid sequence encoded by the nucleotide sequence of SEQ ID NO:
- 44 or a variant polypeptide thereof, wherein the variant has at least 72% sequence identity or more with the sequence set out in SEQ ID NO:
- 43.
A polynucleotide which comprises:
- (a) the nucleotide sequence set out in SEQ ID NO:
- 44;
- (b) a nucleotide sequence which hybridizes selectively with a polynucleotide being the reverse complement of SEQ ID NO:
- 44;
- (c) a nucleotide sequence having at least 74% sequence identity or more with the nucleotide sequence of SEQ ID NO:
- 44;
- (d) a fragment of a nucleotide sequence as defined in (a), (b) or (c) which is at least about 100 nucleotides in length;
- (e) a sequence which is degenerate as a result of the genetic code to a sequence as defined in any one of (a), (b), (c) or (d);
- (f) a nucleotide sequence which is the reverse complement of a nucleotide sequence as defined in (a), (b), (c), (d) or (e).
A process for the in-situ detoxification of a lignocellulose hydrolysate containing furanic compounds, such as preferably one or more of HMF, Furfurylalcohol, Furfural and/or Furoic acid, with a suitable host microorganism, the process comprising a host microorganism from the family of Burkholderiaceae and contacting the lignocellulose hydrolysate with the host microorganism under conditions facilitating the expression of one or more of a Furoyl-CoA dehydrogenase, a Furoyl-CoA synthetase, a 2-oxoglutaroyl-CoA hydrolase, a 2,5-furan-dicarboxylic acid decarboxylase 1, a 2,5-furan-dicarboxylic acid decarboxylase 2 and a HMF/furfural oxidoreductase from the host microorganism from the family of Burkholderiaceae to convert furanic compounds, such as HMF, Furfurylalcohol, Furfural and/or Furoic acid, to non-toxic components to obtain a detoxified lignocellulose hydrolysate.
A process for the production of Furoyl-CoA dehydrogenase and/or Furoyl-CoA synthetase and/or 2-oxoglutaroyl-CoA hydrolase and/or 2,5-furan-dicarboxylic acid decarboxylase 1 and/or 2,5-furan-dicarboxylic acid decarboxylase 2 and/or a HMF/furfural oxidoreductase which are at least 45% identical to those expressed by Cupriavidus basilensis HMF14 DSM 22875, comprising
- (a) culturing a microorganism in a nutrient medium containing carbon and nitrogen sources and inorganic salts;
- and
- (b) isolating the enzymes produced from the microorganism.
A process for the conversion of 5-hydroxymethylfurfural (HMF), 2,5-dihydroxymethyl furan (HMF alcohol), 5-hydroxymethyl-2-furancarboxylic acid (HMF acid) and/or 2,5-furandicarboxylic acid to 2-furoyl CoA, comprising contacting furfuryl alcohol and/or furfural with a furoyl-CoA dehydrogenase, a furoyl-CoA synthetase, a 2-oxoglutaroyl-CoA hydrolase, and a 2,5-furan-dicarboxylic acid decarboxylase catalyst in the presence of one or more coenzyme cofactor(s).
A method for the biocatalytic production of biofuels such as ethanol in a suitable host microorganism of the enzymatic pathway of Cupriavidus basilensis strain DSM 22875, under conditions facilitating the expression of the activity of the enzymes in the enzymatic pathway.
A process for the production of second-generation biofuels from a lignocelluloses-containing material, which process comprises
- a) pretreating and hydrolysis of the lignocellulose-containing material to prepare a lignocellulosic hydrolysate comprising one or more furanic compounds and one or more fermentable sugars;
- b) in-situ detoxification of the lignocellulose hydrolysate comprising contacting the lignocellulose hydrolysate with a host microorganism under conditions facilitating the expression of one or more of a Furoyl-CoA dehydrogenase, a Furoyl-CoA synthetase, a 2-oxoglutaroyl-CoA hydrolase, a 2,5-furan-dicarboxylic acid decarboxylase 1, a 2,5-furan-dicarboxylic acid decarboxylase 2 and/or a HMF/furfural oxidoreductase from the host microorganism to convert the one or more furanic compounds to non-toxic components to obtain a detoxified lignocellulose hydrolysate;
- c) conversion of the fermentable sugars from the lignocellulose hydrolysate or the detoxified lignocelluloses hydrolysate to produce a biofuel.
Dependencies are inferred from the claim text.