Chemistry: molecular biology and microbiology – Treatment of micro-organisms or enzymes with electrical or... – Modification of viruses
Patent
1986-06-26
1988-07-12
Wiseman, Thomas G.
Chemistry: molecular biology and microbiology
Treatment of micro-organisms or enzymes with electrical or...
Modification of viruses
435 68, 435 70, 435253, 435317, 435320, 935 27, 935 40, 935 41, 935 43, 935 48, 935 60, 935 73, C12P 2100, C12P 2102, C12N 1500, C12N 120
Patent
active
047570130
ABSTRACT:
Methods and compositions are provided for regulated expression and secretion of polypeptides in transformed bacterial hosts. A novel class of plasmid cloning vehicles includes a DNA sequence coding for the desired polypeptide (or an insertion site therefor) linked for transcriptional expression in reading phase with four functional fragments derived from the lipoprotein gene of E. coli. The plasmids also include a DNA fragment coding for the signal peptide of the ompA protein of E. coli, positioned such that the desired polypeptide is expressed with the ompA signal peptide at its amino terminus, thereby allowing efficient secretion across the cytoplasmic membrane. The plasmids further include a DNA sequence coding for a specific segment of the E. coli lac promoter-operator, which is positioned in the proper orientation for transcriptional expression of the desired polypeptide, as well as a separate functional E. coli lacI gene coding for the associated repressor molecule which can interact with the lac promoter-operator to prevent transcription therefrom. Expression of the desired polypeptide is under the control of both the lipoprotein promoter and the lac promoter-operator, although transcription from either promoter is normally blocked by the repressor molecule. However, the repressor can be selectively inactivated by means of an inducer molecule to permit transcriptional expression of the desired polypeptide from both promoters. The methods utilize such plasmids to introduce genetic capability into micro-organisms for the production of normally secreted proteins, such as medically or commercially useful hormones, enzymes, immunogenic proteins, or intermediates therefor, but only in the presence of an appropriate inducer.
REFERENCES:
Ghrayeb, J, H. Kimura, M. Takahara, H. Hsiung, Y. Masui and M. Inouye, The EMBO Journal 3(10): 2437-2442, 1984.
Movva, R. N, K. Nakamura and M. Inouye, Journal of Biological Chemistry 255 (1):27-29, 1980.
Matteucci et al, Bio/Technology, vol. 4, pp. 51-55, Jan. 1986.
Gray et al, Gene, vol. 39, pp. 247-254, 1985.
Oka et al, Proc. Natl. Acad. Sci. USA, vol. 82, pp. 7212-7216, Nov. 1985.
Nakamura et al, The EMBO Journal, vol. 1, pp. 771-775, Jun. 1982.
Nakamura et al, Journal of Molecular and Applied Genetics, vol. 1, pp. 289-299, Jun. 1982.
Masui et al, Bio/Technology, vol. 2, pp. 81-85, Jan., 1984.
Masui et al, "Multipurpose Expression Cloning Vehicles in Escherichia Coli," published as Chap. 2 of Experimental Manipulation of Gene Expression (Inouye, M., ed.), Academic Press 1983.
Inouye Masayori
Masui Yoshihiro
Teskin Robin Lyn
The Research Foundation of State University of New York
Wiseman Thomas G.
LandOfFree
Cloning vehicles for polypeptide expression in microbial hosts does not yet have a rating. At this time, there are no reviews or comments for this patent.
If you have personal experience with Cloning vehicles for polypeptide expression in microbial hosts, we encourage you to share that experience with our LandOfFree.com community. Your opinion is very important and Cloning vehicles for polypeptide expression in microbial hosts will most certainly appreciate the feedback.
Profile ID: LFUS-PAI-O-664231