64 results about "Wnt protein secretion" patented technology

Methods are provided for producing recombinant proteins by utilizing expression vectors carrying nucleic acids encoding the proteins, and secretory signal sequences to direct the secretion of the proteins to the periplasm or extracellular medium. Expression vectors which encode a fusion protein comprising a carrier protein and the protein are also provided, as are host cells transformed with the expression vectors.

The present invention provides methods of isolating of leader peptides capable of directing export of heterologous proteins from the bacterial cytoplasm. The methods rely on the screening of libraries of putative leader peptides or of leader peptide mutants for sequences that allow rapid export and thus can rescue a short-lived reporter protein from degradation in the cytoplasm. The mutant leader peptides identified herein are shown to confer significantly higher steady state levels of export not only for short-lived reporter protein but also for other stable, long-lived proteins. These leader peptides can be used to direct or enhance protein secretion. The present invention further discloses methods for the export of cytoplasmically folded protein via the Tat pathway. Proteins having disulfide bonds are first folded within the cytoplasm in suitable oxidizing mutant strains. Such cytoplasmically pre-folded proteins containing disulfide bonds are then exported via the Tat pathway.

The invention discloses a signal peptide for mediating protein secretion expression and belongs to the field of protein engineering and gene engineering. By modifying known signal peptides PelB and OmpA, the N terminal of the signal peptide contains the first three amino acid residues MKK of the signal peptide OmpA, the C terminal of the signal peptide contains the last three amino acid residues AQA of the signal peptide OmpA, and a middle hydrophobic structure is the hydrophobic sequence LLPTAAAGLLLLAAQP of the signal peptide PelB. After the signal peptide capable of effectively improving protein secretion efficiency is used, the secretion expression capability of a strain on target protein is obviously improved; the extracellular enzyme activity of phospholipase D of target protein can be improved by 2 times, the extracellular enzyme production capability of the modified strain is significantly improved, and industrialized application is better facilitated.

Described herein are methods to enhance protein secretion in a host cell. In preferred embodiment, the host cell is a gram-positive microorganism such as a Bacillus. In another preferred embodiment, the host cell is a gram-negative microorganism. Preferably the gram-negative microorganism is an Escherichia coli or a member of the genus Pantoaea. Protein secretion may be enhanced by the overexpression of protein components of the Tat pathway. Alternatively, secretion of foreign proteins can be selectively enhanced by forming a chimeric polypeptide comprising a tat signal sequence and the protein of interest. In a preferred embodiment, the tat signal sequence is selected from phoD or LipA.

Provided is a process for producing a protein with post-translational modification in a cell-free protein synthesis system having a higher protein synthetic activity and glycosylation capability. The process of the present invention comprises preparing a cell extract from cultured cells of an immortalized mammalian cell line that has enhanced protein secreting activity, and adding an mRNA encoding a glycoprotein to the cell extract.

The invention provides a method of making and secreting a non-secreted protein. The method includes expressing the protein from a nucleic acid construct which includes: (a) a mammary epithelial specific promoter; (b) a milk protein specific signal sequence which can direct the secretion of a protein; (c) optionally, a sequence which encodes a sufficient portion of the amino terminal coding region of a secreted protein to allow secretion in the milk of a transgenic mammal, of the non-secreted protein; and (d) a sequence which encodes a non-secreted protein, wherein elements (a), (b), optionally (c), and (d) are preferably operatively linked in the order recited. Both glutamic acid decarboxylase (GAD) and myelin basic protein (MBP), which are cytoplasmic proteins, have been produced by the methods of the present invention. The invention also provides methods for treating diabetes and multiple sclerosis using proteins produced by the methods of the present invention.

The present invention provides compositions and methods for modulating the secretion of transcriptionally regulated proteins from recombinant cells. More particularly, the present invention provides compositions and methods for accelerating the secretion dynamics of human insulin from recombinant cells using nonsense mediated mRNA decay of the preproinsulin mRNA transcript.

The present invention relates to a novel protein secretion factor, a vector including a nucleic acid sequence encoding the protein secretion factor, and a transformed cell into which the vector is introduced. The invention also relates to a method of producing a target protein using the transformed cell including the vector.

Presented herein are methods of evaluating cellular activity by: placing a cell population on an area; assaying for a dynamic behavior of the cell population as a function of time; identifying cell(s)of interest based on the dynamic behavior; characterizing a molecular profile of the cell(s); and correlating the obtained information. The assayed dynamic behavior can include cellular activation, cellular inhibition, cellular interaction, protein expression, protein secretion, cellular proliferation, changes in cellular morphology, motility, cell death, cell cytotoxicity, cell lysis, and combinations thereof. Sensors associated with the area may be utilized to facilitate assaying. Molecular profiles of the cell(s) can then be characterized by various methods, such as DNA analysis, RNA analysis, and protein analysis. The dynamic behavior and molecular profile can then be correlated for various purposes, such as predicting clinical outcome of a treatment, screening cells, facilitating a treatment, diagnosing a disease, and monitoring cellular activity.

An improved mammalian expression vector system which allows: (1) highly expressing exogenous proteins in host mammalian cells; (2) rapidly and efficiently screening recombinant stable cell lines expressing the gene of interest; (3) maintain sustainable expression and prevent gene silencing; and (4) effectively secreting proteins into media in some of cases. The entire expression vector system includes optimized promoters and core promoters, the use of special internal ribosome entry sites, integration of the bacterial backbone into the mammalian expression unit, multiple choices of selection markers, artificial matrix attachment region elements, effective secreting lead sequences and their 5′ and 3′ UTRs, and proper combinations of these expression elements.

The present teachings provide methods for increasing protein secretion, e.g., chymosin in filamentous fungi by co-expressing certain chaperone(s) and / or foldase(s). The present teachings also provide filamentous fungi containing certain chaperone(s) and / or foldase(s) and a protein of interest for increased secretion.

The invention relates to a novel strategy for achieving secretion expression of heterogeneous protein in bacillus subtilis. Non-classical secretion proteins Eno, YceD and YvgN are taken as transport signals to achieve the secretion expression of heterogeneous protein BgaB, but under the guiding of YceD, the secretion quantity of BgaB is low. Under the condition of PdhA and KatA, all through very high quantity of fused protein in a cell is detected, activity of corresponding protein and BgaB outside the cell is not detected, and that PdhA and KatA can not achieve the secretion expression of BgaB is proved. When GapA is taken as a fused protein, fused protein inside the cell and fused protein outside the cell are both not detected, and possibly, the expression of fused protein in the cell does not exist. The experimental method provides novel possibility for secretion expression of heterogeneous protein in bacillus subtilis.

The present invention relates to methods for increasing the secretion of a protein of interest (POI) from a eukaryotic cell comprising co-expression of a POI and of at least one protein that enhances protein secretion, said enhancing protein being selected from the group consisting of BMH2, BFR2, C0G6, C0Y1, CUP5, IMH 1, KIN2, SEC31, SSA4 and SSE1. The invention further relates to a yeast promoter sequence, in particular to a promoter sequence of the PET9 gene of P. pastoris, having, under comparable conditions, an increased promoter activity relative to a promoter sequence of the GAP protein. The invention further relates to an expression vector comprising such a promoter sequence and to the use of such an expression vector for expression of a POI in a host cell. The invention further relates to new yeast promoter sequences of genes from P. pastoris, which are useful for expression of a POI in yeast.

The present invention discloses a medical material with heterogeneous coating layer and its preparation method. It is composed of medical matrix and heterogeneous coating layer coated on the surface of said medical matrix, the described heterogeneous coating layer is formed by distributing zinc, magnesium or strontium contained calcium phosphate or calcium carbonate coacervate whose gain size is 200nm-20 microns into the calcium phosphate coating layer containing manganese and containing fluorine, in which the surface of calcium phosphate coating layer containing manganese or containing fluorine has high surface potential adsorption function, it is favorable for promoting adsorption of cell on its surface, the zinc, magnesium or strontium contained calcium phosphate or calcium carbonate coacervate has ion-releasing function, it can utilize the mode of releasing beneficial ions to promote cytogenic protein secretion and promote mineralization, etc.

The present invention relates to the improved production of proteins, preferably enzymes such as lipases. In particular, the invention relates to a mutated signal peptide and nucleotide sequence encoding said signal peptide that results in an increased protein secretion. Further, the invention relates to a mutated promoter that results in an increased protein expression. It was surprisingly found that the combination of the mutated signal peptide and the mutated promoter act synergistically to result in an about 100-fold increased protein production. Nucleic acid molecules, expression vectors and host cells comprising the mutated signal peptide, mutated promoter, or the combination thereof are also encompassed by the invention. Finally, the invention relates to methods and uses of such nucleic acid molecules, expression vectors and host cells for protein preparation.

Microfluidic “organ-on-a-chip” devices have been developed with the aim to replicate human tissues in vitro. However, there is no option to quantitatively monitor biological processes that take place within the chip, over time. Destructive methods in order to analyze, tissue formation, gene expression, protein secretion etc. require the harvest of the “tissue” at a certain time point. Described herein are methods and compositions for non-destructive molecular imaging methods and systems in order to quantitatively monitor specific biological processes, over time, within the chip, without the need to harvest.

The present disclosure provides mutant cells for the secretion of proteins and for the degradation of lignocellulosic biomass. Methods for the use of these cells are also provided. Specifically, the utility of combined genetic deletions of β-glucosidases and the catabolite repressor gene creA / cre-1 for protein secretion in fungal and yeast cells is disclosed.

An expression vector is disclosed which contains a promoter DNA; a DNA encoding a peptide having a defined amino acid sequence and having secretion signal activity; and a DNA encoding an intended protein or a cloning site for insertion of the DNA encoding an intended protein. An expression vector is also disclosed which contains a promoter DNA; a DNA encoding any peptide having a defined amino acid sequence and having secretion signal activity; a DNA encoding an intended protein or a cloning site for insertion of the DNA encoding an intended protein; and a DNA encoding an anchor domain. The peptide having secretion signal activity allows for secretory production and cell surface display of a protein with high activity, in yeast. According to the present invention, a secretion signal peptide is provided which stably has higher secretion activity ability It is also an object of the present invention to provide a secretion signal peptide that stably has higher secretion ability than that of a conventionally used secretion signal peptide in secretory production and cell surface display of a protein.

The invention opened a constructing method of the secretory donor plasmid for the Bombyx Baculovirus expression system. It is based on the donor plasmid of pFastBacHTa,b,c and the Baculovirus transforming carrier pAcGP67-B. The plasmid provided the condition for getting the recombined protein.

The present invention provides a method for efficiently producing an industrially useful protein in coryneform bacteria, and more particularly, a method for efficiently producing a protein for which secretion was difficult with conventional protein secretion pathways. In particular, the present invention provides a method for efficiently producing heterologous proteins comprising: culturing coryneform bacteria containing an genetic construction containing a promoter sequence which functions in coryneform bacteria, a nucleic acid sequence encoding a Tat system-dependent signal peptide region, and a nucleic acid sequence encoding a heterologous protein, in the direction from 5'-end to 3'-end, and secretory producing the heterologous protein by coryneform bacteria.

The present invention relates to: an expression cassette which has an increased protein secretion in yeast compared to wild-type and which comprises a polynucleotide for coding a translational fusion partner (TFP) and a polynucleotide for coding a protein selected from the group consisting of: TrXynII, TrBx1, TrEGL2, PaCEL1, PaCel2, TeCBH1, NfCBH1, HgCBH1, CtCBH1, ClCBH2, and CfCex1; an expression vector comprising the same; a transformant in which the expression vector is introduced into a host cell; and a complex strain comprising two or more of the transformants. In addition, the present invention relates to a method for producing hemicellulase, endoglucanase, and exoglucanase, or a method for producing bioethanol, comprising a step for culturing the transformants. Furthermore, the present invention relates to a cellulase cocktail comprising endoglucanase and exoglucanase produced by the method and beta-glucosidase, and a method for saccharifying biomass using the cocktail. Moreover, the prevent invention relates to a method for producing bioenergy or useful biochemicals from cellulosic biomass, using a single strain which produces the cellulase, or two or more strains in combination.

The bacterial plant pathogen Pseudomonas syringae injects effector proteins into host cells via a type III protein secretion system to cause disease. The invention relates to the discovery that the type III effector HopU1 is a mono-ADP-ribosyltransferase (ADP-RT) and suppresses plant innate immunity. The HopU1 substrates in Arabidopsis thaliana extracts were RNA-binding proteins that possess RNA-recognition motifs (RRMs). A. thaliana knock-out lines defective in the glycine-rich RNA-binding protein AtGRP7, a HopU1 substrate, were more susceptible than wild type plants to P. syringae. The ADP-ribosylation of AtGRP7 by HopU1 required two arginines within the RRM. The invention provides novel methods for the modulation of the innate immune response of a plant to a biotic stress, including methods for enhancing or suppressing the innate immune response of the plant.

The present invention relates to nucleic acid sequences and amino acid sequences for Bacillus subtilis disulfide bond isomerases, Dsb1 and Dsb2 and methods for increasing the secretion of heterologous and homologous proteins in Gram-positive microorganisms.

The present invention relates to a method of increasing resistance against fungal pathogens of the family Phacosporaceae plants and / or plant cells. This is achieved for instance by increasing the expression of a hydrophobin protein or fragment thereof in a plant, plant part and / or plant cell in comparison to wild type plants, wild type plant parts and / or wild type plant cells. In the transgenic plants hydrophobin can be expressed as a fusion protein to facilitate and / or enhance expression. Furthermore, the hydrophobin protein can be expressed including a secretion signal sequence which mediates secretion of the protein into the apoplast and / or into the cuticule.

The present teachings provide methods for increasing protein secretion, e.g., chymosin in filamentous fungi by co-expressing certain chaperone(s) and / or foldase(s). The present teachings also provide filamentous fungi containing certain chaperone(s) and / or foldase(s) and a protein of interest for increased secretion.

The present invention discloses a method for preparing medical implantation material with amorphous oxide ion delayed-release surface layer. Said method includes the following steps: (1), under the ice water bath environment mixing phosphate or partially-esterified phosphoric mixture with fluorinated compound, then adding calcium salt which can be dissolved in liquid alcohol, heating and refluxing said mixed solution, adding salt containing trace element to form sol; and (2), soaking metal base body in said sol, then adopting impregnation pulling process or rotary coating process to coat the surface of said metal base body, drying and making heat-treatment at 500-750 deg.C.