36results about How to "Improve secretion efficiency" patented technology

The present invention provides novel purified and isolated nucleic acid sequences encoding procoagulant-active FVIII proteins. The nucleic acid sequences of the present invention encode amino acid sequences corresponding to known human FVIII sequences, wherein residue Phe309 is mutated. The nucleic acid sequences of the present invention also encode amino acid sequences corresponding to known human FVIII sequences, wherein the APC cleavage sites, Arg336 and Ile562, are mutated. The nucleic acid sequences of the present invention further encode amino acid sequences corresponding to known human FVIII sequences, wherein the B-domain is deleted, the von Willebrand factor binding site is deleted, a thrombin cleavage site is mutated, an amino acid sequence spacer is inserted between the A2- and A3-domains. Methods of producing the FVIII proteins of the invention, nucleotide sequences encoding such proteins, pharmaceutical compositions containing the nucleotide sequences or proteins, as well as methods of treating patients suffering from hemophilia, are also provided.

Provided is a method for improving secretion efficiency of a recombinant foreign protein in a yeast expression system. The method includes transforming a yeast host with a recombinant foreign gene construct containing a galactose-inducible promoter, a secretion signal sequence and a gene encoding the foreign protein to construct a transformed yeast strain; and culturing the transformed yeast strain under the condition that the activity of the galactose-inducible promoter is controlled. Improved secretion efficiency of the foreign protein can be achieved by decreasing overexpression-induced insoluble precipitation of the recombinant foreign protein suffered by a conventional galactose-inducible promoter-based yeast expression system, via appropriate control of a level of galactose functioning as an inducer of the galactose-inducible promoter in cells. Due to improved secretion efficiency of the recombinant foreign protein, present invention makes a contribution to improvement in productivity of recombinant foreign proteins in the yeast expression system and reduction in production costs.

A method of identifying nucleotide sequences coding for signal peptides in lactic acid bacteria, using a DNA molecule comprising a transposon including a promoterless reporter gene from which DNA molecule a region between the LR and the reporter gene is deleted and the DNA molecule comprises a DNA sequence coding for a secretion reporter molecule. By deleting the region between the LR and the reporter gene, stop codons in-frame with the secretion reporter molecule is removed which upon transposition permits translational fusions from upstream the LR.

A method for improving the secretion rate of exogenous protein expressed by silkworm cells, designing and synthesizing an RNAi interference fragment for interfering with the BiP gene in silkworm BmN cells, transfecting BmN cells for 24 hours and then continuing to transfect the expression vector carrying the foreign gene, after 72 hours That is, the secretion rate of the foreign protein can be significantly increased. This method is simple and easy to operate, and can effectively improve the secretion efficiency of exogenous proteins expressed in silkworm cells. It can provide a new idea to improve the expression of membrane proteins by using BiP etc. as molecular targets, and is conducive to in-depth understanding of baculovirus Expression system, and provide a reference for the mechanism research of signal peptide involved in protein secretion.

The invention provides a method for improving recombinant protein secretion efficiency, which is characterized in that a secretion signal sequence SS in the homeotic allotype domain of the mouse transcription factor Engrailed 2 is fused to the N-terminal of the recombinant protein; wherein the amino acid sequence of the secretion signal sequence SS is that: Ala Gln Glu Leu Ser Leu Asn Glu Ser Gln. The invention has the advantages that: the secretion efficiency of human catalase using the invention is improved to 2.3 times than the normal secretion efficiency guided by bovine Beta-casein signal peptide and provides an effective solution for improving recombinant protein secretion efficiency in industrial production.

The invention discloses a signal peptide capable of improving secretion efficiency. A nucleotide sequence is shown in 1) or 2): 1) the nucleotide sequence is shown as SEQ ID NO.1; 2) after transforming the SEQ ID NO.1 sequence, the nucleotide sequence still performs the function of signal peptide and the secretion efficiency of the nucleotide sequence does not have fundamental change. The invention provides a signal peptide capable of improving the secretion efficiency, the secretion capability of strains can be remarkably improved after the signal peptide is used, and the enzyme activity of asparaginase can be improved by 1.5 times. The enzyme production capability of the transformed strains is remarkably improved, and the transformed strains are more suitable for industrial application, the production cost can be lowered, and the production efficiency can be improved.

The invention relates to a method for the production and non-covalent surface display of antibodies and derived fragments as well as molecule libraries based thereon on the surface of S. cerevisiae cells. The non-covalent manner of the surface display renders possible the selection of specific variants by means of high throughput screening and the subsequent switchable secretion of the selected binding molecule into the culture supernatant for biochemical characterization.

The invention discloses a signal peptide for optimizing high-level secretory expression of keratinase Ker and application of the signal peptide and belongs to the fields of genetic engineering and enzyme engineering. The signal peptide is characterized in that 46 secretory signal peptides from bacillus subtilis are preliminarily screened and 6 secretory signal peptides capable of obviously improving the secretion volume of the keratinase in the 46 secretory signal peptides are directionally transformed, so the signal peptide for optimizing the high-level secretory expression of keratinase is obtained. An N terminal of the signal peptide is provided with first nine amino acid residues of a signal peptide AbnA; an H terminal of the signal peptide is provided with a hydrophobic sequence of a signal peptide YfkN, and a C terminal of the signal peptide is provided with last four amino acid residues of a signal peptide PhoB. The signal peptide disclosed by the invention is fused at the N terminal of the keratinase Ker, so the secretion efficiency of the keratinase of recombinant bacillus subtilis is remarkably improved, and the enzyme activity of extracellular keratinase of the bacillus subtilis is improved by 3.39 times. The extracellular keratinase producing capability of a transformed strain is remarkably improved, the industrial application is better facilitated, the production cost can be reduced, and the production efficiency is improved.