13 results about "Colibiogen" patented technology

The invention discloses a polypeptide asparagine ligase VdiPAL1 and an application of the polypeptide asparagine ligase VdiPAL1. The polypeptide asparagine ligase VdiPAL1 is newly found from viola plants, 12 mg / L zymogen protein is obtained through expression of an escherichia coli prokaryotic system, and compared with homologous enzymes VyPAL2 and VdiPAL1 which are widely applied at present, the polypeptide asparagine ligase VdiPAL1 has the advantages of high expression, high catalytic efficiency and substrate specificity and has good development and application potential. The invention also shows the value of the VdiPAL1 for guiding homologous enzyme modification, and particularly discloses the mutant protein VyOPT of the polypeptide asparagine ligase VyPAL2 and a coding cDNA (complementary deoxyribonucleic acid) sequence of the mutant protein VyOPT. The invention also discloses a mutant VyOPT, which comprises five mutations obtained by modification guided by the VdiPAL1. Compared with the expression quantity of the natural VyPAL2, which is about 0.1 mg / L, the expression quantity of the VyOPT reaches 12 mg / L.

The invention belongs to the technical field of biology, and relates to a preparation method and application of a shark nano antibody (VNAR) targeting HER2 (human epidermal growth factor receptor 2). The shark nano antibody is named as VNAR-G4, and the amino acid sequence of the shark nano antibody is as shown in SEQ ID No. 10. The molecular weight of the VNAR-G4 is about 15kDa, which is far less than that of the traditional monoclonal antibody (-150kDa); the antibody has an extended CDR3 ring structure, can be specifically combined with hidden epitopes (such as a dimerization interface) of HER2, and overcomes the epitope recognition limitation of a traditional antibody; the structural integrity and binding activity can still be maintained under an extreme pH condition, and the catalyst is suitable for a harsh in-vivo environment; experiments prove that the kit has high affinity (KD < = nM level) and strong specificity with HER2 antigen, and has no cross reaction; soluble expression is realized by adopting an escherichia coli prokaryotic expression system, the process is simple, the cost is low, and large-scale production is easy; flexible genetic engineering modification (such as construction of bispecific antibodies or drug conjugates) is supported; the VNAR-G4 can be used for effectively inhibiting the activity of an HER2 signal channel and remarkably inhibiting the proliferation of HER2 positive tumor cells in vitro and in an animal model; and a novel targeted therapy tool is provided for treating HER2-driven malignant tumors such as breast cancer and gastric cancer.

The invention discloses a fusion protein for degrading a cell membrane surface protein and application thereof. The fusion protein is obtained by connecting a diphtheria toxin receptor binding domain with a cell surface target protein ligand molecule through a connecting peptide. The fusion protein disclosed by the invention can be subjected to prokaryotic expression through simple escherichia coli, is efficient and high in yield, has relatively high uniformity, makes up for the defect of a technology of synthesizing LYTAC through complex coupling to a certain extent, can degrade 65% of human epidermal growth factor receptor 2 (Her2) after being treated for 16 hours, and has a good application prospect. And reference and selection are provided for application of the fusion protein in drug development.

This invention discloses a strain that produces high levels of L-homoserine, its construction method, and its applications, belonging to the field of amino acid fermentation. This invention provides a strain for efficient fermentation production of L-homoserine, its construction method, and its applications. This strain utilizes a cofactor homogenization scheme to introduce a heterologous NADH-dependent dehydrogenase to replace the original NADPH-dependent dehydrogenase in *E. coli*, thereby removing cofactor limitations, knocking out the main byproduct pathway, enhancing precursor supply, and strengthening efflux protein expression. Fed-batch fermentation in a fermenter achieves a yield as high as 155 g / L.

The application provides a high-activity recombinant heat-sensitive UDG mutant, which is characterized by that the amino acid sequence is shown in any one of SEQ ID NO: 1, 2 or 5. Its prokaryotic expression vector is also disclosed. The application carries out point mutation on the basis of wild-type heat-sensitive UDG (psychrophilic marine bacterium), the mutant can significantly improve the heat sensitivity of UDG, and improve the digestion capacity of the enzyme on the U-containing template in vitro. By fusing SUMO protein at the N terminal of the vector, the recombinant protein can be obtained in large quantities through supernatant expression in the prokaryotic expression system of Escherichia coli and affinity purification. The recombinant protein can have higher compatibility for reaction systems such as qPCR. The best effect is obtained by UDGm-1, UDGm-2 and UDGm-5.

The invention provides a chicken mycoplasma synoviae subunit vaccine, a preparation method, detection and application thereof. The chicken mycoplasma synoviae VY930 protein expressed on the basis of an escherichia coli prokaryotic expression system has good immunogenicity, and a subunit vaccine containing the protein is prepared from the chicken mycoplasma synoviae VY930 protein so as to effectively prevent and control chicken mycoplasma synoviae infection. Animal challenge protection tests prove that the subunit vaccine has immune protection potential equivalent to or slightly superior to that of an inactivated vaccine, and clinical symptoms caused by the mycoplasma synoviae can be remarkably relieved. The subunit vaccine disclosed by the invention is simple and convenient in preparation process and low in cost, can adapt to a large-scale production mode, and has the potential of preventing and controlling clinical infection of the mycoplasma synoviae and promoting a population purification process. The invention further provides a monoclonal antibody aiming at the protein, the monoclonal antibody can be used as an index for judging whether immunization is carried out or not, meanwhile, the immune protection evaluation criterion after chicken mycoplasma synoviae foot pad challenge is refined, and a meticulous measurement standard is provided for follow-up vaccine research and development.

The application provides a Mycoplasma synoviae subunit vaccine, a preparation method, detection and application thereof. The Mycoplasma synoviae VY930 protein expressed based on an Escherichia coli prokaryotic expression system has good immunogenicity, and a subunit vaccine containing the protein is prepared, so as to effectively prevent and control Mycoplasma synoviae infection. It is proved by animal challenge protection test that the subunit vaccine has equivalent or slightly better immunoprotective potential than the inactivated vaccine, and can significantly reduce the clinical symptoms caused by Mycoplasma synoviae. The subunit vaccine has the potential of preventing and controlling Mycoplasma synoviae clinical infection and promoting the purification process of the population. The application also provides a monoclonal antibody for the above protein, which can be used as an index for judging whether immunization is performed or not, and further refines the immune protection evaluation criteria after footpad challenge of Mycoplasma synoviae, so as to provide a detailed measurement standard for subsequent vaccine research and development.

The invention provides a preparation method of L-cysteine hydrochloride, and relates to the field of chemical processes. According to the preparation method of the L-cysteine hydrochloride, escherichia coli BL21 is adopted as an original host strain, and site-directed mutagenesis is performed on serine transacetylase in the escherichia coli original strain through feedback inhibition mutation, threonine synthetase gene deletion and serine transacetylase gene overexpression, so that the L-cysteine hydrochloride is obtained. By overexpressing the O-acetylserine sulfydryl lyase gene, promoting the combination of hydrogen sulfide and OAS to generate L-cysteine and removing the feedback inhibition of threonine, the method overcomes the defects that a large amount of hydrochloric acid is used and acidic waste is discharged in the preparation of a traditional process, reduces the discharge of waste acid by 80% or more, is renewable in raw materials, meets the requirements of green chemistry, and is suitable for industrial production. The future trend of a green and environment-friendly preparation process is achieved.

The present invention relates to the field of biotechnology, and in particular to a preparation method and application of a recombinant human long-acting interleukin-22 binding protein. The amino acid sequence of the recombinant human long-acting interleukin-22 binding protein is shown in SEQ ID No.1. The present invention utilizes an Escherichia coli (E.coli) prokaryotic expression system to prepare a method for IL-22BP recombinant protein with high biological activity, by designing a novel fusion protein, so that the sequence encoding human IL-22BP is connected to the sequence encoding ABD, constructed as an IL-22BP-ABD fusion gene, induced to be expressed in Escherichia coli, after separating the inclusion body protein, the target protein is obtained by denaturation, purification, and renaturation, and the biological activity of IL-22BP-ABD is verified by in vivo and in vitro experiments.

The application discloses a method for improving soluble expression and functional characteristics of potato protein Patatin based on consensus sequence design, and belongs to the technical field of biotechnology.The application first adopts consensus sequence design to obtain potato ConPatatin protein which can be expressed in a completely soluble form in Escherichia coli, and effectively improves the problem of low yield caused by poor solubility of existing Patatin protein in recombinant expression in the Escherichia coli.The application adopts the Escherichia coli prokaryotic expression system to express the ConPatatin protein in a soluble form, and the protein yield can reach 200 mg / L.The ConPatatin protein prepared in the application has similar protein structure and higher functional activity to the natural extracted Patatin protein, is beneficial to the application of the potato Patatin protein in the fields of food and biological medicine, and lays a beneficial foundation for development of the potato Patatin protein.

The application of the invention is a divisional application of 2022111274353. The invention provides a high-activity recombinant thermosensitive UDG mutant. The high-activity recombinant thermosensitive UDG mutant is characterized in that the amino acid sequence of the mutant is shown as SEQ ID NO: 2 or 5. The invention also discloses a prokaryotic expression vector. According to the invention, point mutation is carried out on the basis of wild-type thermosensitive UDG (psychothermosensitive UDG), the thermal sensitivity of the UDG can be obviously improved by the mutant, and the digestion ability of the enzyme on a U-containing template in vitro is improved. The SUMO protein is subjected to fusion expression at the N end of the vector, and the recombinant protein can be obtained in quantity through affinity purification after being subjected to supernatant expression in an escherichia coli prokaryotic expression system. And the method has higher compatibility with reaction systems such as qPCR (quantitative polymerase chain reaction) and the like. Wherein the optimal effect is UDGm-1; uDGm-2 (UDGm-2); and UDGm-5.

The invention discloses a method for improving soluble expression and functional characteristics of potato protein Patatin based on consensus sequence design, and belongs to the technical field of biology. According to the invention, the consensus sequence design is adopted for the first time to obtain the potato ConPatatin protein which can be expressed in escherichia coli in a completely soluble form, and the problem of low yield caused by poor recombinant expression solubility of the existing Patatin protein in escherichia coli is effectively improved. An escherichia coli prokaryotic expression system is adopted, the ConPatatin protein in a soluble form is expressed, and the protein yield can reach 200 mg / L. The ConPatatin protein prepared by the invention has a similar protein structure and a higher functional activity with naturally extracted Patatin protein, is beneficial to the application of the potato Patatin protein in the fields of food and biological medicine, and lays a beneficial foundation for the development of the potato Patatin protein.

The invention discloses a nano antibody with PEDV (porcine epidemic diarrhea virus) neutralizing activity and application of the nano antibody. The invention relates to the technical field of biological medicines, and discloses a nano antibody NbH2 with PEDV (porcine epidemic diarrhea virus) neutralizing activity, which is obtained from a natural nano antibody library of camelidae animals through an escherichia coli prokaryotic expression system and a phage display technology on the basis of S1 protein of the PEDV, is a VHH antibody, has an amino acid sequence shown as SEQ ID NO.1, and has a nucleotide sequence shown as SEQ ID NO.2. A nucleotide sequence for coding the nano-antibody is shown as SEQ ID NO.2. The nano-antibody has good PEDV neutralizing activity, has a 100% neutralizing effect on PEDV epidemic strains at a dilution degree of 1: 2, can be used for PEDV detection, can also be used as a therapeutic antibody, can be used for preparing PED therapeutic drugs, has important significance in development of antiviral drugs, and has wide application prospects. Meanwhile, a direction is pointed out for exploration of PEDV vaccine targets.