Method for the manufacture of lantibiotics

A technology of lantibiotics and cell culture, applied in peptide preparation methods, chemical instruments and methods, organic chemistry, etc., can solve problems such as complex preparation methods of lantibiotics

Inactive Publication Date: 2009-03-11
BOEHRINGER LNGELHEIM VETMEDICA GMBH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] In summary, the preparation methods known in the art for lantibiotics are complex and require several purification steps

Method used

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  • Method for the manufacture of lantibiotics
  • Method for the manufacture of lantibiotics
  • Method for the manufacture of lantibiotics

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0059] Fermentation of gallidermin

[0060] Freezing culture (WCC):

[0061] A vial of frozen stock (Zellbank BO-002; Kampagne 1940122004033, Reference 35 / 3 / 18) of Staphylococcus gallinarum (Tü 3928; DSM4616) was used to sterile inoculate the agar plates. The agar culture was incubated at 37°C for 1 day, followed by 1 day at 4°C (with Parafilm - but this step is not necessary). This culture was used to inoculate the first generation of seeds. For each production, a new vial must be used.

[0062] plate culture medium

[0063] Yeast Extract Hy-Yest 455 5g / L Kerry Bioscience Glucose x H2O 1g / L Cerestar or Merck NaCl 5g / L Fluka or Merck Bacto Agar 12g / L Bekton Dickinson h 2 o Deionization pH 7.3 (before sterilization)

[0064] Preculture or first generation seeds (2 shake flasks; 500ml):

[0065] One cultured plate (approximately 0.5 x 0.5 cm) was aseptically transferred into two 500 ml baffled shake flasks (a ba...

Embodiment 2

[0086] Purification of gallidermin

[0087] Starting from the culture supernatant of the fermentation process as described in Example 1, gallidermin was purified as follows:

[0088] Method A: Precipitation with Ammonium Sulfate

[0089] After fermentation, the cells were removed by centrifugation, and the cell-free supernatant (pH 6. 7) to precipitate the product.

[0090] In a separate experiment, precipitation with various concentrations of (saturated) ammonium sulfate was tested.

[0091] % Saturation of Ammonium Sulfate Yield(%) 20 9 30 31 40 69 50 76 60 74

[0092] The suspension was stirred slowly for another 30 minutes, then the precipitate was collected by filtration. The precipitate was dissolved in 40% ethanol, 1% aqueous acetic acid (1 / 4 of the starting volume). The yield of the product was 87% with a purity of 93.6 area % as determined by analytical HPLC.

[0093] The purity of the product can be further improved to >...

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Abstract

The present invention relates to the manufacture of antibiotic compounds of the class known as lantibiotics. Preferably, the present invention relates to the purification of those lantibiotics.

Description

field of invention [0001] The present invention relates to the preparation of biomolecules. In particular, the present invention relates to the preparation of antibiotic compounds known to be classified as lantibiotics. Preferably, the invention relates to the purification of those lantibiotics. Background of the invention [0002] Lantibiotics are a class of small peptide antibiotics characterized by the presence of unique, bridged thioether amino acids, namely lanthione and 3-methyllanthione. Members of this class include subtilisin, nisin, epidermin, gallidermin (which is a leucine-6 ​​variant of epidermin), pep5, angiotensin-converting enzyme inhibitory peptide, Ro 09-0198, cinnamycin, and duramycin white. Pep5, epidermin, and Gallierdermin are all naturally produced by microorganisms of the Staphylococcus genus. [0003] The fermentation of Gallidermin-producing strain Staphylococcus gallinarum (S.gallinarium) Tü 3928 (DSM4616) is described in detail in EP-342 486, ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K1/30C07K1/14
CPCC07K14/31C07K1/30C07K1/00C07K1/14
Inventor 休伯特·米尔纳马丁·A·福尔杰安德烈亚斯·沃纳尤维·吉尔里克库尔特·艾尔克劳斯·海因茨曼尼古拉斯·M·肖费边·怀尔
Owner BOEHRINGER LNGELHEIM VETMEDICA GMBH
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