Coryneform bacterium transformant and process for producing dicarboxylic acids using the same

• US 20070087423A1
• Filed: 07/28/2004
• Published: 04/19/2007
• Est. Priority Date: 07/29/2003
• Status: Active Grant

• Current Assignee: Research Institute Of Innovative Technology For The Earth
• Original Assignee: Research Institute Of Innovative Technology For The Earth
• Inventors: Yukawa, Hideaki, Inui, Masayuki, Ikenaga, Yuko, Nakata, Kaori, Okino, Shohei, Murakami, Shikiko
• Granted Patent: US 7,368,268 B2
• Time in Patent Office: Days
• US Class Current: 435/145
• CPC Class Codes: C12N 1/205 Bacterial isolates
• CPC Class Codes: C12P 7/46 Dicarboxylic acids having four or less carbon atoms, e.g. fumaric acid, maleic acid
• CPC Class Codes: C12R 2001/15 Corynebacterium

Abstract

The present invention provides an aerobic coryneform bacterium transformant in which a lactate dehydrogenase gene is disrupted, and a pyruvate carboxylase gene is recombined so as to be highly expressed by a genetic engineering method. The aerobic coryneform bacterium transformant of the present invention can produce dicarboxylic acids from saccharides at a high production rate.

Claims

1. An aerobic coryneform bacterial transformant, wherein a lactate dehydrogenase gene is disrupted and a pyruvate carboxylase gene is recombined so as to be highly expressed, and wherein said aerobic coryneform bacterium is one selected from the group consisting of Corynebacterium, Brevibacterium, Arthrobacter, Mycobacterium and Micrococcus.

2. The aerobic coryneform bacterial transformant according to claim 1, wherein the Corynebacterium is any one of bacteria selected from Corynebacterium glutamicum R, Corynebacterium glutamicum ATCC 13032, Corynebacterium glutamicum ATCC 13058, Corynebacterium glutamicum ATCC 13059, Corynebacterium glutamicum ATCC 13060, Corynebacterium glutamicum ATCC 13232, Corynebacterium glutamicum ATCC 13286, Corynebacterium glutamicum ATCC 13287, Corynebacterium glutamicum ATCC 13655, Corynebacterium glutamicum ATCC 13745, Corynebacterium glutamicum ATCC 13746, Corynebacterium glutamicum ATCC 13761, Corynebacterium glutamicum ATCC 14020 and Corynebacterium glutamicum ATCC 31831.

3. The aerobic coryneform bacterial transformant according to claim 1, wherein the Brevibacterium is any one of bacteria selected from Brevibacterium lactofermentum ATCC 13869, Brevibacterium flavum MJ-233, Brevibacterium flavum MJ-233AB-41 and Brevibacterium ammoniagenes ATCC 6872.

4. The aerobic coryneform bacterial transformant according to claim 1, wherein the Arthrobacter is any one of bacteria selected from Arthrobacter globiformis ATCC 8010, Arthrobacter globiformis ATCC 4336, Arthrobacter globiformis ATCC 21056, Arthrobacter globiformis ATCC 31250, Arthrobacter globiformis ATCC 31738 and Arthrobacter globiformis ATCC 35698.

5. The aerobic coryneform bacterial transformant according to claim 1, wherein the Mycobacterium is Mycobacterium bovis ATCC 19210 or Mycobacterium bovis ATCC 27289.

6. The aerobic coryneform bacterial transformant according to claim 1, wherein the Micrococcus is any one of bacteria selected from Micrococcus freudenreichii No. 239, Micrococcus luteus No. 240, Micrococcus ureae IAM 1010 and Micrococcus roseus IFO 3764.

7. The aerobic coryneform bacterial transformant according to claim 2, wherein the Corynebacterium is Corynebacterium glutamicum R.

8. The aerobic coryneform bacterial transformant according to claim 1, wherein a lactate dehydrogenase gene is disrupted in such a manner that said gene is interrupted, or a part or the whole region of the gene is deleted by a method selected from a homologous recombination method, a transposon insertion method and a mutagen introduction method, and an expression function for the lactate dehydrogenase activity is lost.

9. The aerobic coryneform bacterial transformant according to claim 1, which is Corynebacterium glutamicum R ldh⁻

/pCRB1-PC or Corynebacterium glutamicum R ldh⁻

/pCRB1-PC-FRD.

10. A process for producing dicarboxylic acids, which comprises reacting a bacterium and saccharides in a reaction solution under the reducing state containing carbonate or bicarbonate ions or carbon dioxide gas, and collecting dicarboxylic acids produced in the reaction solution, wherein the bacterium is the aerobic coryneform bacterial transformant as defined in claim 1.

11. The process for producing dicarboxylic acids according to claim 10, wherein an oxidation-reduction potential of the reaction solution under the reducing state is −

200 millivolts to −

500 millivolts.

12. The process for producing dicarboxylic acids according to claim 10, wherein the dicarboxylic acid is selected from succinic acid, fumaric acid and malic acid.