Production of human proinsulin using a novel vector system
US5460954A · kind A · utility
Assignee
Inventors
Key dates
| Filing date | Sep 30, 1992 |
| Grant date | Oct 24, 1995 |
| Priority date | — |
| Expiry date | Sep 30, 2012 |
Classification
- Technology area (CPC C)Chemistry; Metallurgy
- CPC primaryC07K2319/75
- WIPO fieldBiotechnology
- WIPO sectorChemistry
Abstract
The specification describes a process for producing human proinsulin in Escherichia coli (E. coli) using gene manipulation technology. The process can provide for human proinsulin in high yields by a novel expression vector having strong regulatory elements of an insulin gene and a stable recombinant gene product. The expression vector of the present invention is characterized in that: 1) it has an 11 amino acid leader peptide containing six threonines in order to ensure an intracellular stability of proinsulin fusion protein, 2) it contains two copies of a DNA expression cassette each comprising a strong lambda P.sub.R promoter, a lac ribosome binding site, a proinsulin gene with a 17 amino acid leader peptide sequence containing a DNA sequence encoding (Thr).sub.6, and a strong fd phage transcription terminator (combination of phage fd terminator and translation stop codon), etc. successively ligated, 3) it has an ampicillin resistance gene, 4) it can be very stably retained within a cultured cell, and 5) there are a number of these expression vectors in E. coli by which the expression can be significantly increased. Human insulin is prepared from the proinsulin fusion protein by…
Source: USPTO / EPO open patent data. Objective bibliographic and citation counts.