Genes encoding exopolysaccharide production
OwnerMattheos Koffas, James M. Odom +3inventors
InventorsMattheos Koffas, James M. Odom, Siqun Wang, Tao Wang, Rich W. Ye
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Abstract
Genes have been isolated from a Methylomonas sp encoding elements of the exopolysaccharide biosynthetic pathway. The genes and gene products are the first isolated from an organisms capable of utilizing single carbon (C1) substrates as energy sources. The genes are useful for engineering other C1 utilizing microorganisms to make altered levels of exopolysaccharide which is used in a variety of commercial applications.
Claims34
An isolated nucleic acid molecule encoding a Methylomonas sp exopolysaccharide biosynthetic enzyme, selected from the group consisting of:
- (a) an isolated nucleic acid molecule encoding the amino acid sequence selected from the group consisting of SEQ ID NOs:
- 2, 4, 6, 8, 10, 12, 14, 16, and 18;
- (b) an isolated nucleic acid molecule that hybridizes with (a) under the following hybridization conditions:
- 0.1×SSC, 0.1% SDS, 65° C. and washed with 2×SSC, 0.1% SDS followed by 0.1×SSC, 0.1% SDS;
- and
- (c) an isolated nucleic acid molecule that is complementary to (a) or (b).
An isolated nucleic acid molecule comprising a first nucleotide sequence encoding a polypeptide of at least 293 amino acids that has at least 58% identity based on the Smith-Waterman method of alignment when compared to a polypeptide having the sequence as set forth in SEQ ID NO:
- 2, or a second nucleotide sequence comprising the complement of the first nucleotide sequence.
An isolated nucleic acid molecule comprising a first nucleotide sequence encoding a polypeptide of at least 473 amino acids that has at least 36% identity based on the Smith-Waterman method of alignment when compared to a polypeptide having the sequence as set forth in SEQ ID NO:
- 4, or a second nucleotide sequence comprising the complement of the first nucleotide sequence.
An isolated nucleic acid molecule comprising a first nucleotide sequence encoding a polypeptide of at least 366 amino acids that has at least 36% identity based on the Smith-Waterman method of alignment when compared to a polypeptide having the sequence as set forth in SEQ ID NO:
- 6, or a second nucleotide sequence comprising the complement of the first nucleotide sequence.
An isolated nucleic acid molecule comprising a first nucleotide sequence encoding a polypeptide of at least 779 amino acids that has at least 35% identity based on the Smith-Waterman method of alignment when compared to a polypeptide having the sequence as set forth in SEQ ID NO:
- 8, or a second nucleotide sequence comprising the complement of the first nucleotide sequence.
An isolated nucleic acid molecule comprising a first nucleotide sequence encoding a polypeptide of at least 472 amino acids that has at least 23% identity based on the Smith-Waterman method of alignment when compared to a polypeptide having the sequence as set forth in SEQ ID NO:
- 10, or a second nucleotide sequence comprising the complement of the first nucleotide sequence.
An isolated nucleic acid molecule comprising a first nucleotide sequence encoding a polypeptide of at least 272 amino acids that has at least 28% identity based on the Smith-Waterman method of alignment when compared to a polypeptide having the sequence as set forth in SEQ ID NO:
- 12, or a second nucleotide sequence comprising the complement of the first nucleotide sequence.
An isolated nucleic acid molecule comprising a first nucleotide sequence encoding a polypeptide of at least 284 amino acids that has at least 21% identity based on the Smith-Waterman method of alignment when compared to a polypeptide having the sequence as set forth in SEQ ID NO:
- 14, or a second nucleotide sequence comprising the complement of the first nucleotide sequence.
An isolated nucleic acid molecule comprising a first nucleotide sequence encoding a polypeptide of at least 398 amino acids that has at least 26% identity based on the Smith-Waterman method of alignment when compared to a polypeptide having the sequence as set forth in SEQ ID NO:
- 16, or a second nucleotide sequence comprising the complement of the first nucleotide sequence.
An isolated nucleic acid molecule comprising a first nucleotide sequence encoding a polypeptide of at least 317 amino acids that has at least 51% identity based on the Smith-Waterman method of alignment when compared to a polypeptide having the sequence as set forth in SEQ ID NO:
- 18, or a second nucleotide sequence comprising the complement of the first nucleotide sequence.
A method of obtaining a nucleic acid molecule encoding a Methylomonas sp exopolysaccharide biosynthetic enzyme comprising:
- (a) synthesizing an at least one oligonucleotide primer corresponding to a portion of the sequence selected from the group consisting of SEQ ID NOs:
- 1, 3, 5, 7, 9, 11, 13, 15, and 17;
- and
- (b) amplifying an insert present in a cloning vector using the oligonucleotide primer of step (a);
- wherein the amplified insert encodes a portion of an amino acid sequence encoding a Methylomonas sp exopolysaccharide biosynthetic enzyme.
A method for the production of exopolysaccharide comprising:
- contacting a transformed host cell under suitable growth conditions with an effective amount of a carbon source whereby exopolysaccharide is produced, said transformed host cell comprising a set of nucleic acid molecules encoding SEQ ID NOs:
- 2, 4, 6, 8, 10, 12, 14, 16, and 18;
- under the control of suitable regulatory sequences.
A method of regulating exopolysaccharide biosynthesis in an organism comprising, over-expressing at least one isoprenoid gene selected from the group consisting of SEQ ID NO:
- 1, 3, 5, 7, 9, 11, 13, 15, and 17 in an organism such that the exopolysaccharide biosynthesis is altered in the organism.
Dependencies are inferred from the claim text.