Method for producing L-leucine and related DNA and microorganism

A leucine and amino acid technology, applied in the field of DNA, can solve the problem of no L-leucine-resistant Escherichia bacteria and no L-leucine-resistant L-leucine production ability relationship issues

Inactive Publication Date: 2001-01-17
AJINOMOTO CO INC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] However, bacteria of the genus Escherichia that are resistant to L-leucine have not been found, nor has the relationship between L-leucine resistance and L-leucine production ability been found.

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0102] Example 1: Obtaining DNA encoding mutant IPMS

[0103] (1) Obtaining L-leucine-producing strains

[0104] The L-leucine-producing E. coli strain was obtained from the standard laboratory wild-type strain E. coli K-12 by the selection method described below. Escherichia coli K-12 strain was treated with a mutagen solution containing 0.2 mg / ml N-methyl-N-nitro-N-nitrosoguanidine at 37°C for 30 minutes. Then washed twice with sodium chloride solution (0.8%), plated on M9 agar medium containing 1 mg / ml L-leucine analogue 4-azoleucine. The colony that appeared after incubation at 37°C for 5 days was picked to detect its ability to produce L-leucine. Strain No. 9. No. 68, No. 58, No. 55 and No. 15 produces leucine.

[0105] (2) Obtaining the leuA gene from the L-leucine producing strain

[0106] Obtained L-leucine-producing mutant No. The leuA gene of 55 and wild-type strain E. coli K-12 was cloned by in vivo cloning using defective phage Mu d5005 (Groisman, E.A. et...

Embodiment 2

[0113] Example 2: Production of L-leucine by transformants

[0114]Escherichia coli C600 (leu) (Appleyard R.K., Genetics, 39, 440-452 (1954)) used in the production of L-leucine strain No. 9. No. 68, No. 58, No. 55 and No. 15 and P1 phage grown in Escherichia coli K-12 were transduced. Obtain Leu'transducers and test their ability to produce L-leucine. The data are shown in Table 3.

[0115] Donor strain

[0116] *L-leucine production was determined after 48 hours of incubation at 32°C. Data for leucine production represent the average of 10 transductants for each strain.

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PUM

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Abstract

A method for producing L-leucine, comprising the steps of: culturing a bacterium which is transformed with a DNA coding for an alpha-isopropylmalate synthase densensitized in feedback inhibition by L-leucine, in a culture medium to produce and accumulate L-leucine in the medium, and recovering L-leucine from the medium.

Description

Background of the invention [0001] The present invention relates to a DNA encoding a mutant α-isopropylmalate synthase. The present invention also relates to an L-leucine-producing microorganism having a mutant α-isopropylmalate synthase, and a method for producing L-leucine using the microorganism. L-leucine is an essential amino acid, which can be used as food or feed nutritional additives, medical treatment, pharmaceutical or chemical industry reagents or materials, or growth factors for the production of lysine and other amino acids. [0002] In the past, L-leucine was mainly produced by a fermentation method using L-leucine-producing microorganisms belonging to the genus Brevibacterium, Corynebacterium, or Serratia, or mutants thereof (Amino acid fermentation, Japanese Society of Science Press , pp 397-422, 1986). [0003] The highest level of L-leucine accumulation was obtained with Brevibacterium flavum VKPM B-2736. The concentration of L-leucine accumulated in the m...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/09C12N1/20C12N1/21C12N9/10C12N9/88C12N15/31C12N15/60C12N15/63C12P13/06C12R1/19
CPCC12N9/1025C12N9/88C12P13/06
Inventor M·M·古斯雅廷纳M·G·伦特斯Y·I·科兹洛夫L·V·伊万诺维斯卡雅E·B·沃罗希洛瓦
Owner AJINOMOTO CO INC
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