Enzymes

Abstract

The present invention relates to enzymes and processes. In particular, there is described a host 3140 cell transformed or transfected with a nucleic acid encoding a bacterial phytase enzyme and variants thereof.

1 - 45 . (canceled) 
     
     
         46 . A non-naturally occurring nucleotide sequence encoding a phytase obtainable from, preferably derived from,  Buttiauxella  species strain P1-29 deposited under accession number NCIMB 41248. 
     
     
         47 . A nucleotide sequence of  claim 46  wherein the nucleotide sequence has the sequence of SEQ ID NO:2 or a nucleotide sequence that is at least 80% identical thereto. 
     
     
         48 . A nucleotide sequence of  claim 46  wherein the nucleotide sequence has the sequence of SEQ ID NO:2. 
     
     
         49 . A nucleotide sequence of  claim 46  wherein the nucleotide sequence is obtainable from the  Buttiauxella  species strain P1-29 deposited under accession number NCIMB 41248 using primers o29-5 and o29-3. 
     
     
         50 . A nucleotide sequence according to  claim 46 , wherein said nucleotide sequence is isolated and/or purified. 
     
     
         51 . A nucleotide sequence according to  claim 46 , encoding a phytase having an activity maximum at around pH 3 to 6, wherein the activity is determinable by incubating said phytase in a solution containing 2 mM phytate, 0.8 mM CaCl 2  in 200 mM sodium acetate buffer at a temperature of 37° C. 
     
     
         52 . A nucleotide sequence according to  claim 46 , encoding a phytase having an activity maximum at about pH 4.5, wherein the activity is determinable by incubating the phytase in a solution containing 2 mM phytate, 0.8 mM CaCl 2  in 200mM sodium acetate buffer at a temperature of 37° C. 
     
     
         53 . A nucleotide sequence according to  claim 46 , encoding a phytase that comprises at least one substitution in at least one of the following positions (numbering according to the numbering in SEQ ID NO:3): K59, T70, A122, D125, T167, H193, F197, T204, T209, A211, S221, I223, S225, K240, A242, D244, A268, S281, A294, N303, I336 or N351. 
     
     
         54 . A nucleotide sequence according to  claim 46 , encoding a phytase that comprises at least one of the following substitutions (numbering according to the numbering in SEQ ID NO: 3):K59E; T17V; K240T; T1671; K240E; D244C; Q289Y;-T209K or F197S. 
     
     
         55 . A nucleotide sequence according to  claim 46 , encoding a phytase that comprises the following substitutions (numbering according to the numbering in SEQ ID NO:3):
 A122T/T167I/F197S/T209K/A211P/K240E/A242S/S281L/Q289Y/A294E/N303K; or   D125E/T167I/H193R/F197S/T204I/T209K/A211P/K240E/A268V/Q289H/A294E/N303K; or   A122T/T167I/H193R/F197S/T204I/T209K/A211P/K240E/A268V/Q289H/A294E/N303K/I336F; or   N70Y/D125/T167I/H193R/F197S/T204I/T209K/A211P/K240E/A268V/Q289H/A294E/N303K.   
     
     
         56 . A nucleotide sequence according to  claim 46 , encoding a phytase that comprises substitutions in the following positions (numbering according to the numbering in SEQ ID NO:3): A122T/F1671/F197S/F209K/A211P/K240E/A242S/S281L/Q289Y/A294E/N303K. 
     
     
         57 . A food or animal feed composition comprising a phytase encoded by the nucleotide sequence of  claim 46 . 
     
     
         58 . The food or animal feed composition according to  claim 57 , produced by spraying the non-naturally occurring phytase in liquid form onto food or animal feed. 
     
     
         59 . The food or animal feed composition according to  claim 57 , produced by mixing the non-naturally occurring phytase as a dry product with food or animal feed. 
     
     
         60 . A non-naturally occurring nucleotide sequence based on the nucleotide sequence encoding a phytase obtainable from, preferably derived from,  Buttiauxella  species strain P1-29 deposited under accession number NCIMB 41248. 
     
     
         61 . A nucleotide sequence of  claim 60  wherein the nucleotide sequence has the sequence of SEQ ID NO:2 or a sequence that is at least 80% identical thereto. 
     
     
         62 . A nucleotide sequence of  claim 60  wherein the nucleotide sequence has the sequence of SEQ ID NO:2. 
     
     
         63 . A nucleotide sequence of  claim 60  wherein the nucleotide sequence is obtainable from the  Buttiauxella  species strain P1-29 deposited under accession number NCIMB 41248 using primers o29-5 and o29-3. 
     
     
         64 . A nucleotide sequence according to  claim 60 , wherein said nucleotide sequence is isolated and/or purified. 
     
     
         65 . A nucleotide sequence according to  claim 60 , encoding a phytase having an activity maximum at around pH 3 to 6, wherein the activity is determined by incubating said phytase in a solution containing 2 mM phytate, 0.8 mM CaCl 2  in 200 mM sodium acetate buffer at a temperature of 7° C. 
     
     
         66 . A nucleotide sequence according to  claim 60 , encoding a phytase having an activity maximum at about pH 4.5, wherein the activity is determined by incubating the phytase in a solution containing 2 mM phytate, 0.8 mM CaCl 2  in 200 mM sodium acetate buffer at a temperature of 37° C. 
     
     
         67 . A nucleotide sequence according to  claim 60 , encoding a phytase that comprises at least one substitution in at least one of the following positions (numbering according to the numbering in SEQ ID NO:3): K59, T70, A122, D125, T167, H193, F197, T204, T209, A211, S221, I223, S225, K240, A242, D244, A268, 5281, A294, N303, I336 or N351. 
     
     
         68 . A nucleotide sequence according to  claim 60 , encoding a phytase that comprises at least one of the following substitutions (numbering according to the numbering in SEQ ID NO: 3): K59E; T167V; K240T; T1671; K240E; D244C; Q289Y;-T209K or F197S. 
     
     
         69 . A nucleotide sequence according to  claim 60 , encoding a phytase that comprises the following substitutions (numbering according to the numbering in SEQ ID NO: 3):
 A122T/T167I/F197S/T209K/A211P/K240E/A242S/S281L/Q289Y/A294E/N303K; or   D125E/T167I/H193R/F197S/T204I/T209K/A211P/K240E/A268V/Q289H/A294E/N303K; or   A122T/T167I/H193R/F197S/T204I/T209K/A211P/K240E/A268V/Q289H/A294E/N303K/I336F or;   N70Y/D125E/T167I/H193R/F197S/T204I/T209K/A211P/K240E/A268V/Q289H/A294E/N303K.   
     
     
         70 . A nucleotide sequence according to  claim 60 , encoding a phytase that comprises substitutions in the following positions (numbering according to the numbering in SEQ ID NO: 3):
 A122T/F1671/F197S/F209K/A211P/K240E/A242S/S281L/Q289Y/A294E/N303K.   
     
     
         71 . A food or animal feed composition comprising a phytase encoded by the nucleotide sequence of  claim 60 . 
     
     
         72 . The food or animal feed composition according to  claim 71 , produced by spraying the non-naturally occurring phytase in liquid form onto food or animal feed. 
     
     
         73 . The food or animal feed composition according to  claim 71 , produced by mixing the non-naturally occurring phytase as a dry product with food or animal feed. 
     
     
         74 . A polypeptide encoded by the non-naturally occurring nucleotide sequence encoding a phytase obtainable from, preferably derived from,  Buttiauxella  species strain P1-29 deposited under accession number NCIMB 41248. 
     
     
         75 . A polypeptide of  claim 74  wherein the encoding nucleotide sequence has the sequence of SEQ ID NO:2 or a sequence that is at least 80% identical thereto. 
     
     
         76 . A polypeptide of  claim 74  wherein the encoding nucleotide sequence has the sequence of SEQ ID NO:2. 
     
     
         77 . A polypeptide of  claim 74  wherein the encoding nucleotide sequence is obtainable from the  Buttiauxella  species strain P1-29 deposited under accession number NCIMB 41248 using primrs o29-5 and o29-3. 
     
     
         78 . A polypeptide of  claim 74  wherein the encoding nucleotide sequence is isolated and/or purified. 
     
     
         79 . A polypeptide of  claim 74  which is a phytase having an activity maximum at around pH 3 to 6, wherein the activity is determined by incubating said phytase in a solution containing 2 mM phytate, 0.8 mM CaCl 2  in 200 mM sodium acetate buffer at a temperature of 37° C. 
     
     
         80 . A polypeptide of  claim 74  which is a phytase having an activity maximum at about pH 4.5, wherein the activity is determined by incubating the phytase in a solution containing 2 mM phytate, 0.8 mM CaCl 2  in 200 mM sodium acetate buffer at a temperature of 37° C. 
     
     
         81 . A polypeptide according to  claim 74 , which is a phytase that comprises at least one substitution in at least one of the following positions (numbering according to the numbering in SEQ ID NO: 3): K59, T70, A122, D125, T167, H193, F197, T204, T209, A211, S221, I223, S225, K240, A242, D244, A268, S281, A294, N303, I336 or N351. 
     
     
         82 . A polypeptide of  claim 74  which is a phytase that comprises at least one of the following substitutions (numbering according to the numbering in SEQ ID NO: 3): K59E; T167V; K240T; T1671; K240E; D244C; Q289Y;-T209K or F197S. 
     
     
         83 . A polypeptide of  claim 74  which is a phytase that comprises the following substitutions (numbering according to the numbering in SEQ ID NO: 3):
 A122T/T167I/F197S/T209K/A211P/K240E/A242S/S281L/Q289Y/A294E/N303K; or 
 D125E/T167I/H193R/F197S/T204I/T209K/A211P/K240E/A268V/Q289H/A294E/N303K; or 
 A122T/T167I/H193R/F197S/T204I/T209K/A211P/K240E/A268V/Q289H/A294E/N303K/I336F or; 
 N70Y/D125E/T167I/H193R/F197S/T204I/T209K/A211P/K240E/A268V/Q289H/A294E/N303K. 
 
     
     
         84 . A Polypeptide of  claim 74  which is a phytase that comprises substitutions in the following positions (numbering according to the numbering in SEQ ID NO: 3): A122T/F1671/F197S/F209K/A211P/K240E/A242S/S281L/Q289Y/A294E/N303K. 
     
     
         85 . A food or animal feed composition comprising a phytase of  claim 74 . 
     
     
         86 . The food or animal feed composition according to  claim 85 , produced by spraying the non-naturally occurring phytase in liquid form onto food or animal feed. 
     
     
         87 . The food or animal feed composition according to  claim 85 , produced by mixing the non-naturally occurring phytase as a dry product with food or animal feed.