Pichia pastoris formate dehydrogenase and uses therefor
First Claim
1. An isolated polynucleotide comprising a nucleotide sequence encoding amino acid sequence SEQ ID NO:
- 5.
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Abstract
This invention relates to a recombinant Pichia pastoris formate dehydrogenase (FDH) enzyme that catalyzes the oxidation of formate to carbon dioxide and the simultaneous reduction of nicotinamide adenine dinucleotide (NAD+) to its reduced form (NADH). Also related are isolated nucleic acids encoding P. pastoris FDH polypeptides, and fragments and variants thereof, as well as vectors and host cells comprising these nucleic acids. Further related are isolated, recombinant P. pastoris FDH polypeptides, and fragments and variants thereof, and antibodies that specifically bind to P. pastoris FDH polypeptides, fragments, or variants. The invention also relates to methods of obtaining isolated P. pastoris FDH nucleic acids, polypeptides, and antibodies, and methods of using P. pastoris FDH in various reactions for industrial or pharmaceutical applications.
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Citations
15 Claims
- 1. An isolated polynucleotide comprising a nucleotide sequence encoding amino acid sequence SEQ ID NO:
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2. An isolated polynucleotide comprising a nucleotide sequence encoding the NAD-binding domain of amino acid sequence SEQ ID NO:
- 5, wherein said polynucleotide encodes a polypeotide having formate dehydrogenase activity, and wherein the NAD-binding domain of amino acid sequence SEQ ID NO;
5 is amino acids 117-309 of SEQ ID NO;
5. - View Dependent Claims (12, 13, 15)
- 5, wherein said polynucleotide encodes a polypeotide having formate dehydrogenase activity, and wherein the NAD-binding domain of amino acid sequence SEQ ID NO;
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3. An isolated polynucleotide comprising a nucleotide sequence encoding the catalytic domain of amino acid sequence SEQ ID NO:
- 5, wherein said polynucleotide encodes a polypeptide having formate dehydrogenase activity, and wherein the catalytic domain of amino acid sequence SEQ ID NO;
5 is amino acids 16-115 of SEQ ID NO;
5.
- 5, wherein said polynucleotide encodes a polypeptide having formate dehydrogenase activity, and wherein the catalytic domain of amino acid sequence SEQ ID NO;
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6. An isolated polynucleotide which hybridizes under high stringency conditions to any one of:
- (a) the nucleotide sequence SEQ ID NO;
1;
(b) the nucleotide sequence SEQ ID NO;
3;
(c) the complement of (a);
(d) the complement of (b);
or (e) a nucleotide sequence that differs from (a), (b), (c) or (d) due to degeneracy of the genetic code;
wherein said polynucleotide encodes a polypeptide having formate dehydrogenase activity; and
wherein said high stringency conditions are hybridization in 50% formamide, 5×
Denhardt'"'"'s solution. 5×
SSPE, and 0.2% SDS at 42°
C., followed by washing in 0.1×
SSPE and 0.1% SDS at 65°
C.
- (a) the nucleotide sequence SEQ ID NO;
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9. An isolated polynucleotide comprising the nucleotide sequence encoding a formate dehydrogenase contained in the plasmid in the ATCC deposit designated PTA-3691.
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14. An isolated polynucleotide comprising a nucleotide sequence encoding a polypeptide that shares at least 95% sequence identity with amino acid sequence SEQ ID NO:
- 5, wherein said polynucleotide encodes a polypeptide having formate dehydrogenase activity.
Specification