Oxaloacetate hydrolase deficient fungal host cells
AssigneeNovozymes A/S
InventorsCarsten Mailand Hjort, Henrik Pedersen
Life of this patent
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Abstract
The present invention relates to isolated nucleic acid sequences encoding polypeptides having oxaloacetate hydrolase activity. The invention also relates to nucleic acid constructs, vectors, and host cells comprising the nucleic acid sequences as well as recombinant methods for producing the polypeptides.
Claims49
An isolated nucleic acid sequence encoding a polypeptide having oxaloacetate hydrolase activity, selected from the group consisting of:
- (a) a nucleic acid sequence encoding a polypeptide having an amino acid sequence which has at least 65% identity with amino acids 1 to 341 of SEQ ID NO:
- 2;
- (b) a nucleic acid sequence having at least 65% homology with a nucleotide sequence constituted by nucleotides 1157-1411, 1504-1651 and 1764-2383 of SEQ ID NO:
- 1;
- (c) a nucleic acid sequence which hybridizes under medium stringency conditions with (i) the nucleic acid sequence of SEQ ID NO:
- 1, (ii) the cDNA sequence of SEQ ID NO:
- 1, (iii) a subsequence of (i) or (ii) of at least 100 nucleotides, or (iv) a complementary strand of (i), (ii), or (iii);
- (d) an allelic variant of (a), (b), or (c);
- (e) a subsequence of (a), (b), (c), or (d), wherein the subsequence encodes a polypeptide fragment which has oxaloacetate hydrolase activity;
An isolated nucleic acid sequence produced by (a) hybridizing a DNA under medium stringency conditions with (i) the nucleic acid sequence of SEQ ID NO:
- 1, (ii) the cDNA sequence of SEQ ID NO:
- 1, (iii) a subsequence of (i) or (ii) of at least 100 nucleotides, or (iv) a complementary strand of (i), (ii), or (iii);
- and (b) isolating the nucleic acid sequence.
A substantially pure culture of E. coli DSM 12660 or DSM 12661.
A method for producing a product, comprising
- (a) culturing a mutant of a parent cell, wherein the mutant produces less oxaloacetate hydrolase than the parent cell, wherein the oxaloacetate hydrolase
- (i) has an amino acid sequence that has at least 90% identity with amino acids 1-341 of SEQ ID NO:
- 2;
- (ii) is encoded by a nucleic acid sequence having at least 90% homology with a nucleotide sequence comprising nucleotides 1157-1411, 1504-1651 and 1764-2383 of SEQ ID NO:
- 1;
- (iii) is encoded by a nucleic acid sequence which hybridizes under high stringency conditions with the nucleic acid sequence of SEQ ID NO:
- 1, the cDNA sequence of SEQ ID NO:
- 1, a complementary strand of SEQ ID NO:
- 1, and/or a complementary strand of the cDNA sequence of SEQ ID NO:
- 1, wherein the high stringency conditions are defined as prehybridization and hybridization at 42° C. in 5×SSPE, 0.3% SDS, 200 micrograms/ml sheared and denatured salmon sperm DNA, and 50% formamide, followed by washing three time for 15 minutes using 2×SSC, 0.2% SDS at 65° C.;
- and/or
- (iii) a subsequence of one or both of (i) and (ii), wherein the subsequence encodes a fragment that has oxaloacetate hydrolase activity;
- and
- (b) recovering the polypeptide.
Dependencies are inferred from the claim text.