Transformant for producing 4-hydroxybenzoic acid or salt thereof
The mutant chorismate-pyruvate lyase (A) or (B) as described below is capable of producing 4-hydroxybenzoic acid or a salt thereof with sufficient practical efficiency. (A) A mutant chorismate-pyruvate lyase obtained by replacing the valine at position 80 in a chorismate-pyruvate lyase (ubiC) from Pantoea ananatis consisting of the amino acid sequence of SEQ ID NO: 1 with one or more other amino acids. (B) A mutant chorismate-pyruvate lyase obtained by replacing an amino acid in another chorismate-pyruvate lyase, the amino acid being at a position enzymologically homologous with that of the above valine, with one or mere other amino acids.
1. A transformant obtained by introducing into a coryneform bacterium host cell a DNA which encodes a mutant chorismate-pyruvate lyase, wherein the mutant chorismate-pyruvate lyase is a variant of the polypeptide of SEQ ID NO: 1, wherein said variant has at least 90% sequence identity with the polypeptide of SEQ ID NO: 1, 2, 3, 4, 5, 6, 7, 8 or 9, wherein said variant has chorismate-pyruvate lyase activity, and wherein said variant comprises an alanine, cysteine, threonine, serine, or asparagine at the position corresponding to position 80 of the polypeptide of SEQ ID NO: 1.
2. The transformant of claim 1 , wherein the DNA is
(a) a DNA having 90% or more of identity with any one of SEQ ID NO: 10 to 16, wherein gtc at positions corresponding to positions 240 to 242 in a chorismate-pyruvate lyase gene of Pantoea ananatis consisting of the base sequence of SEQ ID NO: 10, a chorismate-pyruvate lyase gene of Providencia stuartii consisting of the base sequence of SEQ ID NO: 11, a chorismate-pyruvate lyase gene of Providencia rustigianii consisting of the base sequence of SEQ ID NO: 12, a chorismate-pyruvate lyase gene of Providencia sneebia consisting of the base sequence of SEQ ID NO: 13, a chorismate-pyruvate lyase gene of Providencia rettgeri consisting of the base sequence of SEQ ID NO: 14, a chorismate-pyruvate lyase gene of Providencia alcalifaciens consisting of the base sequence of SEQ ID NO: 15, or a chorismate-pyruvate lyase gene of Providencia burhodogranariea consisting of the base sequence of SEQ ID NO: 16 is replaced with a DNA segment which encodes a single amino acid selected from the group consisting of alanine, cysteine, threonine, serine, and asparagine,
(b) a DNA having 90% or more of identity with SEQ ID NO: 17, wherein atc at positions corresponding to positions 237 to 239 in a chorismate-pyruvate lyase gene of Escherichia coli consisting of the base sequence of SEQ ID NO: 17 is replaced with a DNA segment which encodes a single amino acid selected from the group consisting of alanine, cysteine, threonine, serine, and asparagine, or
(c) a DNA having 90% or more of identity with SEQ ID NO: 18, wherein atc at positions corresponding to positions 237 to 239 in a chorismate-pyruvate lyase gene of Cronobacter sakazakii consisting of the base sequence of SEQ ID NO: 18 with a DNA segment which encodes a single amino acid that is different from isoleucine.
3. The transformant of claim 1 , wherein the coryneform bacterium host cell is a Corynebacterium glutamicum.
4. The transformant of claim 3 , wherein the Corynebacterium glutamicum is Corynebacterium glutamicum R deposited under Accession Number FERM BP-18976, ATCC13032, or ATCC13869.
5. A Corynebacterium glutamicum transformant, wherein said transformant is Corynebacterium glutamicum HBA-47 deposited at NITE under (Accession Number BP-01849.
6. A process for producing 4-hydroxybenzoic acid or a salt thereof, which comprises a step of culturing the transformant of claim 1 in a reaction mixture containing at least one starting compound selected from the group consisting of a sugar, a compound that can be metabolized into chorismic acid by the transformant, chorismic acid, and a salt thereof, and a step of recovering 4-hydroxybenzoic acid or a salt thereof from the reaction mixture.
7. The process of claim 6 , wherein the transformant is cultured under aerobic conditions.