Chemistry: molecular biology and microbiology – Micro-organism – tissue cell culture or enzyme using process... – Recombinant dna technique included in method of making a...
Patent
1995-08-22
1998-02-03
Robinson, Douglas W.
Chemistry: molecular biology and microbiology
Micro-organism, tissue cell culture or enzyme using process...
Recombinant dna technique included in method of making a...
43525231, 536 241, C12P 2106, C12N 120, C07H 2104
Patent
active
057143463
DESCRIPTION:
BRIEF SUMMARY
This application is a 371 of PCT/JP94/00269, filed Feb. 22, 1994.
1. Field of Art
The present invention relates to a process for production of human growth hormone (hereinafter sometimes referred to as "hGH") by Bacillus brevis carrying a DNA coding for hGH as a heterologous gene, as well as said DNA and Bacillus brevis carrying said DNA.
2. Background Art
The hGH comprises 191 amino acids, and the amino acid sequence thereof is already known (Bancroft, F. C., Expl. Cell Res., 79, 275-278 (1973), or David V. Goeddel et al., Nature, 281, 544-548 (1979)). Since this substance has growth-stimulating action, action in fat-metabolism or action in sugar-metabolism, for a long time it has been used as a therapeutic agent for dwarfism, and recently it has been noticed as a therapeutic agent for senile dementia.
So far, much recombinant DNA research has been carried out using E. coli, and many heterogeneous proteins have been produced in E. coli. However, in this method, since the heterologous protein produced is intracellularly accumulated, not only are extraction of a desired product from the cells and the purification of the desired product from the extract time- and labor-consuming, but also it is not easy to recover the desired product in its native form.
On the other hand, for a long time, microorganisms belonging to the genus Bacillus have been industrially used as producers for various extracellular enzymes. Among these extracellular enzymes, .alpha.-amylase gene of Bacillus amyloliguefaciens) (I. Palva et al., Gene, 22, 229 (1983)), penicillinase gene of Bacillus licheniformis) (C. Chang et al., Molecular Cloning and Gene Regulation in Bacilli, Academic Press, 659 (1982)), and .alpha.-amylase gene of Bacillus subtilis (H. Yamazaki et al., J. Bacteriol., 156, 327 (1983) have been cloned, and production and secretion of heterogeneous proteins using promoters or signal peptides of the above genes have been reported.
For the production and secretion of heterogeneous proteins by the genus Bacillus, Bacillus subtilis is mainly used as a host. However, since this microorganism produces a large amount of extracellular proteases, there is a tendency for heterogeneous proteins secreted by a recombinant DNA technique to be degraded and the amount accumulated is notably decreased.
On the contrary, Udaka et al. succeeded to produce and secrete .alpha.-amylase (Japanese Unexamined Patent Publication (Kokai) No. 62-201,583;, H. Yamagata et al., J. Bacteriol., 169, 1239 (1987)), and swine pepsinogen (Shigezo Udaka, the abstracts of the 1987 Meeting of the Agricultural Chemical Society of Japan, p 837 to p 838; Norihiro Tsukagoshi, Nippon Nogei Kagaku Kaishi, 61, 68 (1987), using a promoter and signal peptide of MWP (Middle Wall Protein) (H. Yamagata et al., J. Bacteriol., 169, 1239 (1987); Norihiro Tsukagoshi, Nippon Nogei Kagaku Kaishi, 61, 68 (1987)), which is a major extracellular protein of the above-mentioned microorganism, using as a host Bacillus brevis 47 which substantially does not extracellularly produce proteases (Japanese Unexamined Patent Publication (Kokai) Nos. 60-58,074, and 62-201,583; FERMP-7224).
In addition, Udaka et al. succeeded in isolating Bacillus brevis HPD31 (FERMBP-1087) producing no detectable extracellular protease, and in using it as a host so as to produce thermostable .alpha.-amylase (Japanese Unexamined Patent Publication (Kokai) No. 63-56,277; the abstracts of the 1987 Agricultural Chemical Society of Japan), and human EGF (H. Yamagata et al., Proc. Natl. Acad. Sci. USA, 86, 3589 (1989); Japanese Unexamined Patent Publication (Kokai) No. 2-31,682).
The production of hGH using as a host E. coli (David V. Geoddel et al., Nature, 281, 544-548 (1979), Chang C. N. et al., Gene, 55, 189-196 (1987)), Bacillus subtilis (M. Honjo et al., J. Biotech., 6, 191-204 (1987)), yeast (R. Hiramatsu et al., Appl. Environ. Microbiol., 57, 2052-2056 (1991)) or animal cells (Lupker J. H. et al., Gene, 24, 281-287 (1983)) has been reported.
DISCLOSURE OF THE INVENTION
As described above, although it has be
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Hirai Masana
Hoshino Fumihiko
Kajino Tsutomu
Saito Yoko
Udaka Shigezo
Robinson Douglas W.
Sumitomo Pharmaceuticals Company Limited
Wai Thanda
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