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US 2006/0234280 A1Published application
Enyzmes having alpha amylase activity and methods of making and using them
AssigneeVerenium Corporation
InventorsWalter Callen, Toby Richardson, Gerhard Frey, Carl Miller, Martin Kazaoka, Jay Short, Eric Mathur
Life of this patent
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Abstract
The invention relates to alpha amylases and to polynucleotides encoding the alpha amylases, and methods of making and using them. In addition methods of designing new alpha amylases and methods of use thereof are also provided. The alpha amylases have increased activity and stability at increased pH and temperature.
Claims75
An isolated or recombinant polypeptide comprising:
- (a) an amino acid sequence having at least 80% sequence identity to SEQ ID NO:2 over a region of at least about 100 residues, wherein the polypeptide has alpha amylase activity;
- (b) an amino acid sequence encoded by a nucleic acid having at least 80% sequence identity to SEQ ID NO:
- 1 over a region of at least about 100 residues, wherein the polypeptide has alpha amylase activity;
- (c) an amino acid sequence encoded by a nucleic acid that hybridizes under conditions of high stringency to SEQ ID NO:
- 1, wherein the conditions of high stringency comprise a wash step comprising a wash for 30 minutes at room temperature in a solution comprising 150 mM NaCl, 20 mM Tris hydrochloride, pH 7.8, 1 mM Na2EDTA, 0.5% SDS, followed by a 30 minute wash in fresh solution, and the polypeptide has alpha amylase activity;
- (d) an amino acid sequence having at least 70% sequence identity to SEQ ID NO:2, and having at least one or more conservative amino acid substitutions, deletions or insertions, wherein the polypeptide has alpha amylase activity, and optionally the conservative amino acid substitutions comprises:
- substitution of a isoleucine, valine, leucine or methionine for another;
- substitution of arginine for lysine, glutamic acid for aspartic acid or glutamine for asparagine;
- or
- (e) an amino acid sequence comprising (i) a fragment of at least 20 consecutive amino acids of SEQ ID NO:2, or (ii) an amino acid sequence of step (a), (b), (c) or (d), that can generate antibodies which bind specifically to SEQ ID NO:2;
- wherein optionally the sequence identity is determined by analysis with a sequence comparison algorithm or by visual inspection.
8 more elements
An isolated or recombinant polypeptide comprising a sequence as set forth in SEQ ID NO:2, or a functional fragment thereof.
A dry milling process comprising use of a polypeptide having alpha amylase activity for liquefaction of whole ground grain, wherein the polypeptide having alpha amylase activity comprises
- (a) an amino acid sequence encoded by a nucleic acid comprising a sequence
- (i) as set forth in SEQ ID NO:1;
- (ii) having at least 85% sequence identity to SEQ ID NO:1;
- (iii) having a length of at least 200 nucleotides of SEQ ID NO:1 and having at least 95% sequence identity to SEQ ID NO:1;
- (iv) having a length of at least 150 nucleotides of SEQ ID NO:1 and having at least 97% sequence identity to SEQ ID NO:1;
- or
- (v) having a length of at least 75 nucleotides of SEQ ID NO:1 and having at least 99% sequence identity to SEQ ID NO:1.
- (b) an amino acid sequence comprising
- (i) a sequence as set forth in SEQ ID NO:2, or an enzymatically active fragment of SEQ ID NO:2 at least 75, 100 or 150 amino acid residues in length;
- (ii) at least 90% sequence identity to SEQ ID NO:2;
- (iii) an enzymatically active fragment of at least 150 amino acid residues in length and having 95% sequence identity to SEQ ID NO:2;
- or
- (iv) an enzymatically active fragment of at least 75 amino acid residues in length and having 98% sequence identity to SEQ ID NO:2,
- wherein optionally the liquefaction of whole ground grain by the polypeptide is a low temperature enzymatic liquefaction.
11 more elements
A brewing process comprising use of a polypeptide having alpha amylase activity, wherein the polypeptide having alpha amylase activity comprises
- (a) an amino acid sequence encoded by a nucleic acid comprising a sequence
- (i) as set forth in SEQ ID NO:1;
- (ii) having at least 85% sequence identity to SEQ ID NO:1;
- (iii) having a length of at least 200 nucleotides of SEQ ID NO:1 and having at least 95% sequence identity to SEQ ID NO:1;
- (iv) having a length of at least 150 nucleotides of SEQ ID NO:1 and having at least 97% sequence identity to SEQ ID NO:1;
- or
- (v) having a length of at least 75 nucleotides of SEQ ID NO:1 and having at least 99% sequence identity to SEQ ID NO:1.
- (b) an amino acid sequence comprising
- (i) a sequence as set forth in SEQ ID NO:2, or an enzymatically active fragment of SEQ ID NO:2 at least 75, 100 or 150 amino acid residues in length;
- (ii) at least 90% sequence identity to SEQ ID NO:2;
- (iii) an enzymatically active fragment of at least 150 amino acid residues in length and having 95% sequence identity to SEQ ID NO:2;
- or
- (iv) an enzymatically active fragment of at least 75 amino acid residues in length and having 98% sequence identity to SEQ ID NO:2.
10 more elements
A method for textile processing comprising use of a polypeptide having alpha amylase activity, wherein the polypeptide having alpha amylase activity comprises
- (a) an amino acid sequence encoded by a nucleic acid comprising a sequence
- (i) asset forth in SEQ ID NO:1;
- (ii) having at least 85% sequence identity to SEQ ID NO:1;
- (iii) having a length of at least 200 nucleotides of SEQ ID NO:1 and having at least 95% sequence identity to SEQ ID NO:1;
- (iv) having a length of at least 150 nucleotides of SEQ ID NO:1 and having at least 97% sequence identity to SEQ ID NO:1;
- or
- (v) having a length of at least 75 nucleotides of SEQ ID NO:1 and having at least 99% sequence identity to SEQ ID NO:1.
- (b) an amino acid sequence comprising
- (i) a sequence as set forth in SEQ ID NO:2, or an enzymatically active fragment of SEQ ID NO:2 at least 75, 100 or 150 amino acid residues in length;
- (ii) at least 90% sequence identity to SEQ ID NO:2;
- (iii) an enzymatically active fragment of at least 150 amino acid residues in length and having 95% sequence identity to SEQ ID NO:2;
- or
- (iv) an enzymatically active fragment of at least 75 amino acid residues in length and having 98% sequence identity to SEQ ID NO:2.
10 more elements
A method for paper or pulp processing comprising use of a polypeptide having alpha amylase activity, wherein the polypeptide having alpha amylase activity comprises
- (a) an amino acid sequence encoded by a nucleic acid comprising a sequence
- (i) asset forth in SEQ ID NO:1;
- (ii) having at least 85% sequence identity to SEQ ID NO:1;
- (iii) having a length of at least 200 nucleotides of SEQ ID NO:1 and having at least 95% sequence identity to SEQ ID NO:1;
- (iv) having a length of at least 150 nucleotides of SEQ ID NO:1 and having at least 97% sequence identity to SEQ ID NO:1;
- or
- (v) having a length of at least 75 nucleotides of SEQ ID NO:1 and having at least 99% sequence identity to SEQ ID NO:1.
- (b) an amino acid sequence comprising
- (i) a sequence as set forth in SEQ ID NO:2, or an enzymatically active fragment of SEQ ID NO:2 at least 75, 100 or 150 amino acid residues in length;
- (ii) at least 90% sequence identity to SEQ ID NO:2;
- (iii) an enzymatically active fragment of at least 150 amino acid residues in length and having 95% sequence identity to SEQ ID NO:2;
- or
- (iv) an enzymatically active fragment of at least 75 amino acid residues in length and having 98% sequence identity to SEQ ID NO:2.
10 more elements
A method for making a beverage comprising a polypeptide having alpha amylase activity, wherein the polypeptide having alpha amylase activity comprises
- (a) an amino acid sequence encoded by a nucleic acid comprising a sequence
- (i) as set forth in SEQ ID NO:1;
- (ii) having at least 85% sequence identity to SEQ ID NO:1;
- (iii) having a length of at least 200 nucleotides of SEQ ID NO:1 and having at least 95% sequence identity to SEQ ID NO:1;
- (iv) having a length of at least 150 nucleotides of SEQ ID NO:1 and having at least 97% sequence identity to SEQ ID NO:1;
- or
- (v) having a length of at least 75 nucleotides of SEQ ID NO:1 and having at least 99% sequence identity to SEQ ID NO:1.
- (b) an amino acid sequence comprising
- (i) a sequence as set forth in SEQ ID NO:2, or an enzymatically active fragment of SEQ ID NO:2 at least 75, 100 or 150 amino acid residues in length;
- (ii) at least 90% sequence identity to SEQ ID NO:2;
- (iii) an enzymatically active fragment of at least 150 amino acid residues in length and having 95% sequence identity to SEQ ID NO:2;
- or
- (iv) an enzymatically active fragment of at least 75 amino acid residues in length and having 98% sequence identity to SEQ ID NO:2.
10 more elements
Dependencies are inferred from the claim text.