Chemistry: molecular biology and microbiology – Micro-organism – tissue cell culture or enzyme using process... – Recombinant dna technique included in method of making a...
Patent
1997-10-17
1999-11-09
Guzo, David
Chemistry: molecular biology and microbiology
Micro-organism, tissue cell culture or enzyme using process...
Recombinant dna technique included in method of making a...
435 6951, 435 696, 435 697, 43525421, C12P 2100, C12P 2102, C12N 119
Patent
active
059812273
DESCRIPTION:
BRIEF SUMMARY
The invention pertains to the field of recombinant DNA technology and concerns a method for the production of proteins with the aid of genetically engineered yeast cells carrying hybrid vectors comprising the genes for said proteins.
BACKGROUND OF THE INVENTION
Although in genetic engineering numerous protein expression systems for prokaryotic and eukaryotic hosts are already known, there is a continuing demand for novel systems which have advantages over the known systems.
Working on the expression of heterologous proteins in the baker yeast Saccharomyces cerevisiae, it has been commonly observed, that a high-level expression is dependent on many factors, e.g. plasmid stability, plasmid copy number, promoter strength, translation efficiency, low protein degradation.
In this context, one of the very important requisites is the yeast strain which is used for the production.
Recently, quite a number of heterologous proteins have been expressed in different yeast strains after transformation of yeast cells with suitable expression vectors comprising DNA sequences coding for said proteins, like e.g. .alpha.-interferon (Hitzeman et al. Nature (1981), 294, 717-722), tissue-type plasminogen activator (EP-A-143081) or certain desulfatohirudins (EP-A-225633). In many cases, however, the heterologous proteins are not synthesized in pure form, but as a mixture containing partially degraded such as C- or N-terminally shortened proteins. For instance, the expression of human atrial natriuretic peptide (hANP) in yeast resulted in the secretion of two forms of mature hANP differing in their C-terminus (Vlasuk et al. J. Biol. Chem. (1986), 261, 4798-4796). Similar results have been obtained after the expression of epidermal growth factor (EGF) in yeast (George-Nascimento et al. Biochemistry (1988), 27, 797-802) where the secreted expression products were heterologous in that either the last (Arg 53) or the last two amino acids (Leu 52 and Arg 53) were missing and no full-length EGF was produced.
The separation of mixtures containing full-length proteins such as .alpha.-interferon, tissue-type plasminogen activator, inhibitors of tissue-type plasminogen activator, or desulfatohirudins as well as partially degraded like C- or N-terminally shortened derivatives thereof into the individual components and the purification of these components to homogeneity, if these derivatives are biologically active at all, is laborious and time-consuming. Considering the incidental expenses there is a need for improved methods which render possible the economic production of homogenous proteins such as desulfatohirudin in yeast. It is an object of the present invention to provide methods for the production of proteins heterologous to yeast in a homogenous form.
Surprisingly it has been found, that the use of Saccharomyces cerevisiae strain HT393 for the expression of heterologous proteins leads to increased yield of biologically active and undegraded form of the expressed heterologous protein, compared to other Saccharomyces cerevisiae strains that are genetically closely related, e.g., to strain cl3-ABYS-86(DSM 9698) that is genetically closest related.
DESCRIPTION OF THE INVENTION
The present invention concerns a process for the production of a protein heterologous to yeast in a homogenous form characterized in that Saccharomyces cerevisiae strain HT393 (DSM 9697) or a derivative thereof is used for the expression of said heterologous protein.
In a preferred embodiment, the present invention relates to an improved process for the production of a protein heterologous to yeast in a homogenous form comprising culturing Saccharomyces cerevisiae strain HT393(DSM 9697) or a derivative thereof that has been transformed with a hybrid vector comprising a DNA sequence coding for said heterologous protein and isolating said heterologous protein.
A derivative of HT393 is a strain that is derived from HT393 and shows the same properties in respect to the production of heterologous proteins. The use of the inventive strains leads, e.g., to an inc
REFERENCES:
W. Heinemeyer et al. Embo Journal, vol. 10, No. 3, 1991 (pp. 555-562).
T.M. Antalis et al. PNAS, vol. 85, 1988 (pp. 985-989).
Russell et al. The Journal of Biological Chemistry, vol. 258, No. 4, 1983 (pp. 2674-2682).
Price et al. Gene, vol. 55, 1987 (pp. 287-293).
Broker Michael
Meyhack Bernd
Behringwerke AG
Guzo David
McCormack Myra H.
Movartis AG
LandOfFree
Process for the production of proteins does not yet have a rating. At this time, there are no reviews or comments for this patent.
If you have personal experience with Process for the production of proteins, we encourage you to share that experience with our LandOfFree.com community. Your opinion is very important and Process for the production of proteins will most certainly appreciate the feedback.
Profile ID: LFUS-PAI-O-1454243