Thermostable alpha-amylases

Abstract

An isolated polynucleotide comprising an open reading frame encoding a polypeptide having alpha-amylase activity, the polypeptide selected from the group consisting of: a) a polypeptide comprising an amino acid sequence which has at least 70% identity with amino acids 22 to 450 of SEQ ID NO:4; b) a polypeptide comprising an amino acid sequence which has at least 70% identity with the polypeptide encoded by the amylase encoding part of the polynucleotide inserted into a plasmid present in the E. coli host deposited under the Budapest Treaty with DSMZ under accession number DSM 15334; c) a polypeptide encoded by a polynucleotide comprising a nucleotide sequence which has at least 70% identity with the sequence shown from position 68 to 1417 in SEQ ID NO:3; and d) a fragment of (a), (b) or (c) that has alpha-amylase activity.

1 . An isolated polynucleotide comprising an open reading frame encoding a polypeptide having alpha-amylase activity, the polypeptide selected from the group consisting of: 
 a) a polypeptide comprising an amino acid sequence which has at least 70% identity with amino acids 22 to 450 of SEQ ID NO:4;    b) a polypeptide comprising an amino acid sequence which has at least 70% identity with the polypeptide encoded by the amylase encoding part of the polynucleotide inserted into a plasmid present in the  E. coli  host deposited under the Budapest Treaty with DSMZ under accession number DSM 15334;    c) a polypeptide encoded by a polynucleotide comprising a nucleotide sequence which has at least 70% identity with the sequence shown from position 68 to 1417 in SEQ ID NO:3; and    d) a fragment of (a), (b) or (c) that has alpha-amylase activity.    
     
     
         2 . The polynucleotide according to  claim 1 , wherein the polypeptide is an artificial variant comprising an amino acid sequence that has one or more truncation(s), and/or at least one substitution, deletion, and/or insertion of an amino acid as compared to amino acids 22 to 450 of SEQ ID NO:4.  
     
     
         3 . The polynucleotide according to  claim 1 , wherein the polypeptide comprises an amino acid sequence which has at least 70% identity with amino acids 22 to 450 of SEQ ID NO:4.  
     
     
         4 . The polynucleotide according to  claim 1 , wherein the polypeptide comprises the amino acids 22 to 450 of SEQ ID NO:4.  
     
     
         5 . The polynucleotide according to  claim 1 , wherein the polypeptide consists of the amino acids 22 to 450 of SEQ ID NO:4.  
     
     
         6 . The polynucleotide according to  claim 1 , wherein the polypeptide comprises an amino acid sequence which has at least 70% identity with the polypeptide encoded by the amylase encoding part of the nucleotide sequence inserted into a plasmid present in the  E. coli  host deposited under the Budapest Treaty with DSMZ under accession number DSM 15334.  
     
     
         7 . The polynucleotide according to  claim 6 , wherein the polypeptide comprises the amino acid sequence encoded by the amylase encoding part of the nucleotide sequence inserted into a plasmid present in the  E. coli  host deposited under the Budapest Treaty with DSMZ under accession number DSM 15334.  
     
     
         8 . The polynucleotide according to  claim 6  wherein the polypeptide consists of the amino acid sequence encoded by the amylase encoding part of the nucleotide sequence inserted into a plasmid present in the  E. coli  host deposited under the Budapest Treaty with DSMZ under accession number DSM 15334.  
     
     
         9 . The polynucleotide according to  claim 6 , wherein the polypeptide is an artificial variant which comprises an amino acid sequence that has one or more truncation(s), and/or at least one substitution deletion, and/or insertion of an amino acid as compared to the amino acid sequence encoded by the amylase encoding part of the nucleotide sequence inserted into a plasmid present in the  E. coli  host deposited under the Budapest Treaty with DSMZ under accession number DSM 15334.  
     
     
         10 . A nucleic acid construct comprising a polynucleotide as defined in  claim 1  operably linked to one or more control sequences that direct the production of the polypeptide in a suitable host cell.  
     
     
         11 . A recombinant expression vector comprising a nucleic acid construct as defined in  claim 10 .  
     
     
         12 . A recombinant host cell comprising a nucleic acid construct as defined in  claim 10 .  
     
     
         13 . The cell according to  claim 12 , which is a microorganism.  
     
     
         14 . The cell according to  claim 13 , which is a bacterium or a fungus.  
     
     
         15 . The cell according to  claim 14 , which is a Gram-positive bacterium such as  Bacillus subtilis, Bacillus licheniformis, Bacillus lentus, Bacillus brevis, Bacillus stearothermophilus, Bacillus alkalophilus, Bacillus amyloliquefaciens, Bacillus coagulans, Bacillus circulans, Bacillus lautus  or  Bacillus thuringiensis.    
     
     
         16 . The cell according to  claim 14 , which is a protease deficient strain of the fungus  Aspergillus , in particular  A. oryzae.    
     
     
         17 . A method for producing a polypeptide having alpha-amylase activity, the method comprising: 
 (a) cultivating a recombinant host cell as defined in  claim 12  under conditions conducive for production of the polypeptide, and    (b) recovering the polypeptide.    
     
     
         18 . A method of producing an enzymatically modified starch derivative, wherein a polypeptide having alpha-amylase activity produced according to a method as defined in  claim 17  is used for liquefying and/or saccharifying starch.  
     
     
         19 . A method of producing high maltose syrups, wherein a polypeptide having alpha-amylase activity produced according to a method as defined in  claim 17  is used for liquefying starch.  
     
     
         20 . An alcohol production process, wherein a polypeptide having alpha-amylase activity produced according to a method as defined in  claim 17  is used for liquefaction starch in a distillery mash.