Cloning, expression and use of acid phospholipases

摘要

The invention relates to a DNA sequence, which codes for a polypeptide having phospholipase activity essentially without lipase activity, characterized in that the DNA sequence is selected from a) DNA sequences that comprise a nucleotide sequence according to SEQ ID NO: 1, b) DNA sequences that comprise the coding sequence according to SEQ ID NO: 1, c) DNA sequences that code for the protein sequence according to SEQ ID NO: 2, d) DNA sequences that are coded for by the plasmid pPL3940-Topo2.5 with the restriction map according to FIG. 7, which is deposited under accession number DSM 22741, e) DNA sequences that hybridize under stringent conditions with one of the DNA sequences according to a), b), c) or d), f) DNA sequences that are related to the DNA sequences according to a), b), c), d) or e) due to the degeneration of the genetic code, and g) complementary strands to the sequences according to a) to f), wherein the DNA sequence is preferably derived from Aspergillus, and more preferably from Aspergillus fumigatus, and a polypeptide having phospholipase activity essentially without lipase activity selected from a) a polypeptide which is coded for by the coding part of a DNA sequence as defined above, b) a polypeptide having the sequence according to SEQ ID NO: 2 or a sequence derived therefrom, which may be obtained by substitution, addition, deletion of one or more amino acid(s), c) a polypeptide having a sequence that has at least 83% identity with the amino acids 1 to 299 of SEQ ID NO: 2, d) a polypeptide which is coded for by a nucleic acid sequence which hybridizes under stringent conditions with (i) nucleotides 55 to 1106 of SEQ ID NO: 1, (ii) the cDNA sequence contained in nucleotides 55 to 1106 of SEQ ID NO: 1, (iii) a partial sequence of (i) or (ii) composed of at least 100 nucleotides, or (iv) a complementary strand of (i), (ii) or (iii), e) a variant of the polypeptide having SEQ ID NO: 2, comprising a substitution, deletion and/or insertion of one or more amino acid(s), f) allelic variants to amino acid sequences a) to e).

主权项

1. A cDNA sequence that encodes a polypeptide with phospholipase activity essentially without lipase activity comprising a cDNA sequence selected from the group consisting of
a) DNA sequences that comprise a nucleotide sequence according to SEQ ID NO: 1, b) DNA sequences that comprise the coding sequence according to SEQ ID NO: 1, c) DNA sequences that code for the protein sequence according to SEQ ID NO: 2, d) DNA sequences that are coded for by the plasmid pPL3940-Topo2.5 with the restriction map according to FIG. 7, which is deposited under accession number DSM 22741, e) DNA sequences that hybridize under stringent conditions with one of the DNA sequences according to a), b), c) or d), wherein said DNA sequences hybridize at 65° C., 18 h in dextran sulphate solution, subsequent washing of the filter for 30 min, respectively, at first with 6× saline-sodium citrate (SSC), twice 2×SSC, twice 2×SSC, 0.1% SDS, and with 0.2×SSC at 65° C., and f) complementary strands to the sequences according to a) to f), wherein the DNA sequence is derived from Aspergillus.