Genetically modified microorganism and process for production of macrolide compound using the microorganism
OwnerKazuhiro Machida, Yasuhide Aritokuinventors
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Abstract
A genetically modified microorganism capable of directly producing a 16-hydroxylated macrolide compound; and a process for producing a 16-hydroxylated macrolide compound using the microorganism. Specifically, a genetically modified microorganisms having DNA encoding a polypeptide involved in the biosynthesis of a macrolide compound pladienolide and DNA encoding a polypeptide having a pladienolide 16-hydroxylase activity; and a process for producing a 16-hydroxylated macrolide compound comprising the steps of culturing the genetically modified microorganism in a culture medium and collecting a 16-hydroxylated macrolide compound from the culture medium.
Claims21
A recombinant microorganism, which is a microorganism having ability to produce 16-position hydroxyl macrolide compounds represented by the formula (I):
- (wherein R represents a hydrogen atom or hydroxyl group), and, which comprises:
- (a) DNA encoding wholly or partially a polypeptide participating in biosynthesis of the macrolide compounds represented by the formula (II):
- (wherein R represents a hydrogen atom or hydroxyl group);
- and
- (b) DNA encoding wholly or partially a polypeptide having 16-position hydroxylating enzyme activity with respect to the macrolide compounds represented by the above formula (II).
A variant of DNA comprising the continuous nucleotide sequence of nucleotides 1322 to 2548 in the Sequence No. 2 which have 1 or 2 or more modified sites selected from the group consisting of 1) to 6) below:
- 1) a modified site wherein the sequence cgc of nucleotides 1592 to 1594 is modified to a codon encoding an amino acid other than arginine;
- 2) a modified site wherein the sequence atg of nucleotides 1655 to 1657 is modified to a codon encoding an amino acid other than methionine;
- 3) a modified site wherein the sequence cgc of nucleotides 1904 to 1906 is modified to a codon encoding an amino acid other than arginine;
- 4) a modified site wherein the sequence cgc of nucleotides 2027 to 2029 is modified to a codon encoding an amino acid other than arginine;
- 5) a modified site wherein the sequence atc of nucleotides 2054 to 2056 is modified to a codon encoding an amino acid other than isoleucine;
- and
- 6) a modified site wherein the sequence ctg of nucleotides 2528 to 2530 is modified to a codon encoding an amino acid other than leucine.
A variant of DNA comprising the continuous nucleotide sequence of nucleotides 1322 to 2548 in the Sequence No. 2 which have 1 or 2 or more modified sites selected from the group consisting of 1) to 6) below:
- 1) a modified site wherein the sequence cgc of nucleotides 1592 to 1594 is modified to a codon encoding cysteine, methionine or proline;
- 2) a modified site wherein the sequence atg of nucleotides 1655 to 1657 is modified to a codon encoding threonine or serine;
- 3) a modified site wherein the sequence cgc of nucleotides 1904 to 1906 is modified to a codon encoding leucine, isoleucine, proline, tyrosine or phenylalanine;
- 4) a modified site wherein the sequence cgc of nucleotides 2027 to 2029 is modified to a codon encoding leucine, isoleucine or proline;
- 5) a modified site wherein the sequence atc of nucleotides 2054 to 2056 is modified to a codon encoding phenylalanine or valine;
- and
- 6) a modified site wherein the sequence ctg of nucleotides 2528 to 2530 is modified to a codon encoding methionine, cysteine or isoleucine.
A variant of DNA comprising the continuous nucleotide sequence of nucleotides 1322 to 2548 in the Sequence No. 2 which have 1 or 2 or more modified sites selected from the group comprising 1) to 6) below:
- 1) a modified site wherein the sequence cgc of nucleotides 1592 to 1594 is modified to a codon encoding cysteine;
- 2) a modified site wherein the sequence atg of nucleotides 1655 to 1657 is modified to a codon encoding threonine;
- 3) a modified site wherein the sequence cgc of nucleotides 1904 to 1906 is modified to a codon encoding leucine or tyrosine;
- 4) a modified site wherein the sequence cgc of nucleotides 2027 to 2029 is modified to a codon encoding leucine;
- 5) a modified site wherein the sequence atc of nucleotides 2054 to 2056 is modified to a codon encoding phenylalanine;
- and
- 6) a modified site wherein the sequence ctg of nucleotides 2528 to 2530 is modified to a codon encoding methionine.
Dependencies are inferred from the claim text.