Method for secretory production of human growth hormone
AssigneeMitsui Chemicals, Inc.
InventorsMasaru Honjo, Naokazu Naitoh, Hiroshi Uchida, Daisuke Mochizuki, Kazuya Matsumoto
Life of this patent
Estimating the term…
Abstract
A DNA encoding 20K hGH is connected directly to a gene encoding Escherichia coli OppA protein secretion signal, or a modified form thereof, and a DNA encoding signal peptidase 1 to construct a recombinant plasmid, E. coli is transformed by the plasmid and cells of the resulting E. coli transformant strain are cultured for secretory production of the 20K hGH in the E. coli periplasm. This method enables efficient secretory production of 20K hGH and easy isolation and purification of 20K hGH from the periplasm fraction because the level of impure proteins in the E. coli periplasm is low.
Claims29
A recombinant DNA containing a DNA encoding a secretion signal which is more than 60% homologous to the amino acid sequence of the E. coli OppA secretion signal and a DNA encoding a human growth hormone having a molecular weight of about 20,000, wherein the 5′ end of a DNA encoding said human growth hormone having a molecular weight of about 20,000 and the 3′ end of the DNA encoding the secretion signal are operatively linked.
A recombinant plasmid comprising a recombinant DNA in which the 5′ end of a DNA encoding a human growth hormone having a molecular weight of about 20,000 is operatively linked to the 3′ end of a DNA encoding a secretion signal, wherein the DNA encoding a secretion signal is either a DNA encoding E. coli, or Salmonella OppA secretion signal, or a DNA encoding a secretion signal which is different from E. coli or Salmonella OppA secretion signals but is more than 60% homologous to said E. coli OppA secretion signal in its amino acid sequence, and further comprising a DNA encoding E. coli signal peptidase 1.
An E. coli transformant according to claim wherein the E. coli transformant is FERM BP-6290.
A DNA encoding a recombinant secretion signal which is different from E. coli or Salmonella OppA secretion signals but is more than 60% homologous to said E. coli OppA secretion signal in its amino acid sequence, characterized in that a human growth hormone having a molecular weight of about 20,000 can be secreted and produced using a microorganism having a recombinant DNA in which the 3′ end of a DNA encoding said secretion signal is operatively linked to the 5′ end of a DNA encoding the human growth hormone having a molecular weight of about 20,000.
A method according to claim 32 wherein a medium comprising 0.01-10% glycerol, or 0.01-5% succinic acid, or both is used in culturing cells of an E. coli transformant strain.
Dependencies are inferred from the claim text.