Nitrile hydratase
Provided is an improved nitrile hydratase with improved catalytic activity. Also provided are DNA for coding the improved nitrile hydratase, a recombinant vector that contains the DNA, a transformant that contains the recombinant vector, nitrile hydratase acquired from a culture of the transformant, and a method for producing the nitrile hydratase. Also provided is a method for producing an amide compound that uses the culture or a processed product of the culture. The improved nitrile hydratase contains an amino acid sequence represented by SEQ ID NO: 50 (GX 1 X 2 X 3 X 4 DX 5 X 6 R) in a beta subunit, and is characterized in that X 4 is an amino acid selected from a group comprising cysteine, aspartic acid, glutamic acid, histidine, isoleucine, lysine, methionine, asparagine, proline, glutamine, serine and threonine.
1. A modified Rhodococcus bacterial or Nocardia bacterial nitrile hydratase, comprising in the β subunit, an amino-acid sequence as shown in SEQ ID NO: 81:
WEX 1 X 2 X 3 X 4 X 5 X 6 X 7 X 8 X 9 X 10 X 11 X 12 X 13 X 14 X 15 X 16 X 17 X 18 D (SEQ ID NO: 81),
wherein
W is tryptophan,
E is glutamic acid,
D is aspartic acid,
X 1 ˜X 6 , X 8 ˜X 13 and X 15 ˜X 18 each independently indicate any amino-acid residue,
X 7 is an amino acid selected from the group consisting of alanine, valine, aspartic acid, threonine, phenylalanine, isoleucine and methionine, and
X 14 is glycine.
2. The modified Rhodococcus bacterial or Nocardia bacterial nitrile hydratase according to claim 1 , wherein X 1 is G (glycine), X 2 is R (arginine), X 3 is T (threonine), X 4 is L (leucine), X 5 is S (serine), X 6 is I (isoleucine), X 8 is T (threonine), X 9 is W (tryptophan), X 10 is M (methionine), X 11 is H (histidine), X 12 is L (leucine), X 13 is K (lysine), and X 14 is G (glycine) in SEQ ID NO: 81.
3. The modified Rhodococcus bacterial or Nocardia bacterial nitrile hydratase according to claim 1 , further comprising an amino-acid sequence in SEQ ID NO: 82 comprising the amino-acid sequence in SEQ ID NO: 81.
4. A DNA encoding the modified Rhodococcus bacterial or Nocardia bacterial nitrile hydratase according to claim 1 .
5. A recombinant vector, comprising the DNA according to claim 4 .
6. A transformant, comprising the recombinant vector according to claim 5 .
7. A nitrile hydratase collected from a culture obtained by incubating the transformant according to claim 6 .
8. A method for producing a nitrile hydratase, the method comprising:
incubating the transformant according to claim 6 ; and
collecting the nitrile hydratase from the obtained culture.
9. A method for producing an amide compound, the method comprising contacting a nitrile compound with a culture, or a processed product of the culture, obtained by incubating the nitrile hydratase according to claim 1 .