KR960034415A

KR960034415A - Purification Method of Human Interleukin-6 Expressed in Recombinant Escherichia Coli

Info

Publication number: KR960034415A
Application number: KR1019950006994A
Authority: KR
Inventors: 조중명; 최덕영; 소홍섭; 양재영
Original assignee: 성재갑; 주식회사 Lg 화학
Priority date: 1995-03-30
Filing date: 1995-03-30
Publication date: 1996-10-22

Abstract

본 발명은 재조합 대장균에서 발현된 인간 인터류킨-6(interleukin-6,IL-6)를 정제하는 방법에 관한 것으로, 대장균 세포의 파괴 및 불용성 침전물의 회수, 트리톤 X-100을 이용한 불용성 침전물의 세척, 트리톤X-100과 구아니딘 염을 이용한 불용성 침전물의 용해, 트리톤 X-100 존재하의 투석 및 용해성 단백질의 회수, 및 양이온 교환 크로마토그래피를 포함하는 본 발명의 정제 방법은 간단하고 실시가 용이하면서도 고순도의 인간 IL-6를 제조할 수 있다.The present invention relates to a method for purifying human interleukin-6 (IL-6) expressed in recombinant E. coli, the destruction of E. coli cells and recovery of insoluble precipitate, washing of insoluble precipitate using Triton X-100, Purification methods of the present invention, including dissolution of insoluble precipitate using Triton X-100 and guanidine salts, dialysis and recovery of soluble proteins in the presence of Triton X-100, and cation exchange chromatography, are simple and easy to implement IL-6 can be prepared.

Description

Purification Method of Human Interleukin-6 Expressed in Recombinant Escherichia Coli

본 내용은 요부공개 건이므로 전문내용을 수록하지 않았음Since this is an open matter, no full text was included.

제1도는 인간 인터류킨-6(interleukin-6, IL-6) 단백질의 아미노산 서열을 참고로하여 그와 동일한 아미노산 서열을 코드하도록 대장균에서 선호되는 코돈을 사용하여 재설계한 IL-6 유전자의 염기 서열을 나타낸 것이고, 제2도는 중합효소 연쇄 반응을 이용하여 합성한 14개의 올리고뉴클레오티드로부터 IL-6를 포함하는 전체 유전자를 합성하기 위한 연결 전략을 나타낸 것이고, 제3도는 대장균 발현 벡터 pTrpH IL-6의 제작 과정을 나타낸 것이다.Figure 1 refers to the amino acid sequence of human interleukin-6 (IL-6) protein and refers to the nucleotide sequence of the IL-6 gene redesigned using the preferred codon in E. coli to code the same amino acid sequence. FIG. 2 shows a linkage strategy for synthesizing the entire gene including IL-6 from 14 oligonucleotides synthesized using a polymerase chain reaction, and FIG. 3 shows the expression of E. coli expression vector pTrpH IL-6. The production process is shown.

Claims

A) obtaining a precipitate after destroying E. coli cells expressing human interleukin-6, b) washing the precipitate with a nonionic surfactant solution, c) washing the precipitate containing Triton X-100 and guanidine salt Dissolving in a buffer comprising: d) dialysis of the solution in a buffer containing Triton X-100, and e) performing cation exchange chromatography to express human interleukin-6 expressed in recombinant E. coli. How to purify.

The method of claim 1, wherein the nonionic surfactant of step b) is a buffer containing 0.1 to 0.5% of Triton X-100.

The method of claim 1, wherein the buffer of step c) is a buffer containing 0.1 to 0.5% of Triton X-100 and 2 to 6M guanidine salt.

The method of claim 1, wherein the buffer of step d) is a buffer containing 0.1 to 0.5% of Triton X-100.

The method according to claim 1, wherein in step e), Q-sepharose is used as the ion exchange resin and eluted using a buffer containing 0.1 to 0.5% of Triton X-100.

The method of claim 5 wherein a linear gradient of sodium chloride concentration of 0-0.5 M is used during elution.

※ Note: The disclosure is based on the initial application.