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Process for production of cadaverine

A cadaverine and microbial technology, applied in the field of cadaverine preparation, can solve the problems of no increase in cadaverine yield, generation of by-product lysine, increased burden of cadaverine refining, etc., and achieve the goal of reducing load and increasing sugar yield Effect

Active Publication Date: 2012-11-07
TORAY IND INC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, there are many problems to be solved especially in the production method of cadaverine using microorganism culture. By-product lysine is generated (refer to Non-Patent Document 4)
When the by-product lysine is produced, the yield of cadaverine does not increase although the precursor can be produced; in addition, in order to use cadaverine as a raw material for polyamide, high purity of cadaverine is important, but if By-product lysine will increase the burden on cadaverine purification, causing economic problems

Method used

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  • Process for production of cadaverine
  • Process for production of cadaverine
  • Process for production of cadaverine

Examples

Experimental program
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Effect test

Embodiment

[0073] The following examples and comparative examples are listed to describe the present invention in detail. In addition, unless otherwise specified, all culture media, agar media, and culture media used in Examples and Comparative Examples are subjected to general sterilization operations (such as 121 ° C, 30 minutes of high-pressure steam sterilization, or 0.45 μm filter), use in the sterilized state.

[0074] (HPLC analysis method for cadaverine and lysine concentration)

[0075] Column used: CAPCELL PAK C18 (Shiseido)

[0076] Mobile phase: 0.1% (w / w) phosphoric acid aqueous solution: acetonitrile=4.5:5.5

[0077] Detection: UV360nm

[0078] Sample pretreatment: Add 25 μl of 1,4-butanediamine (0.03M) as an internal standard to 25 μl of analysis sample, add 150 μl of sodium bicarbonate (0.075M) and 2,4-dinitrofluorobenzene (0.2M ) in ethanol and incubated at 37°C for 1 hour. 50 μl of the above reaction solution was dissolved in 1 ml of acetonitrile, then centrifuged ...

reference example 1

[0079] Reference Example 1 (Creation of Corynebacterium glutamicum capable of producing lysine)

[0080] In order to produce Corynebacterium glutamicum capable of synthesizing lysine which is a precursor of cadaverine, lysine-producing bacteria were produced by introducing effective mutations into aspartokinase. Corynebacterium glutamicum AK-1 strain (hereinafter referred to as AK-1 strain) was produced by the method described in Apppl. Microbiol. Biotechnol., (2002), 58, p.217-223. The aspartokinase of this strain can relieve the feedback inhibition caused by lysine and threonine. Therefore, lysine can be synthesized by cultivation.

Embodiment 1

[0082] Example 1 (Creation of Corynebacterium glutamicum secreting lysine decarboxylase extracellularly) (Part 1: Utilization of Tat pathway)

[0083] (1) Cloning of HOM gene

[0084] Homoserine dehydrogenase was selected as the locus for introducing the lysine decarboxylase gene. The gene corresponding to the 300 amino acid region from the N-terminal of the HOM gene was cloned. Oligonucleotide primers (SEQ ID NO: 1 and NO: 2) were synthesized with reference to the base sequence of the HOM gene (Accession No: BA000036) registered in the database (GenBank). The genomic DNA solution prepared according to conventional methods from Corynebacterium glutamicum ATCC13032 was used as an amplification template, and 0.2 μl of each was put into a 0.2ml microcentrifuge tube, and various reagents were added: 20 pmol of each primer, pH8.0 Tris hydrochloride buffer (20 mM), potassium chloride (2.5 mM), gelatin (100 μg / ml), each dNTP (50 μM), and LATaq DNA polymerase (2 units) (manufactured...

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Abstract

Disclosed is a process for producing cadaverine, characterized by culturing a microorganism capable of secreting lysine decarboxylase outside cells thereof. According to the process, the by-production of lysine can be prevented, the yield of cadaverine on glucose consumption is improved compared with those in conventional processes, and the load in a purification step for purifying cadaverine as a polyamide raw material can be reduced.

Description

technical field [0001] The present invention relates to the preparation method of cadaverine. In particular, the present invention relates to a method for efficiently producing cadaverine by causing microorganisms to secrete and produce lysine decarboxylase. Background technique [0002] Polyamide (PA) is an important polymer group used as a raw material for a series of special plastics used in the automotive industry, sports industry, and lifestyle industry, and diamine is an important raw material monomer component of the polyamide. Diamines and dicarboxylic acids condense to form various polymers, where the properties of the polymers are determined by the chain lengths of the diamines and dicarboxylic acids. [0003] Generally, diamines are prepared from petroleum-derived materials chemically through an intermediate stage of dicarboxylic acids; or by chemical decarboxylation of amino acids (Non-Patent Document 1). Considering the high price of petroleum, a rapid change ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12P13/00C12N15/09
CPCC07K2319/02C12N9/88C12P13/001
Inventor 耳塚孝须田和美泽井秀树
Owner TORAY IND INC
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