25 results about "Polyhistidine-tag" patented technology

The invention provides fusion protein containing snake venom protein and application of the fusion protein in biosynthesis of stable isotope labeled snake venom protein, and belongs to the technical field of genetic engineering and protein preparation. The fusion protein comprises a 6-poly-histidine tag, a fusion tag, a restriction enzyme cutting site of WELQ protease and snake venom protein which are connected in sequence. According to the fusion tag, the solubility of the snake venom protein fusion protein can be improved; the 6-poly-histidine tag and the fusion tag can be removed through enzyme digestion of WELQ protease, the enzyme digestion removal effect is good, and the natural structure and activity of the snake venom protein can be guaranteed; besides, a culture medium containing a unique nitrogen source or a unique carbon source labeled by an isotope is used for carrying out induction culture on recombinant bacteria for expressing the fusion protein, the fusion protein containing the target snake venom protein is subjected to induction expression, purification, WELQ protease digestion and the like are carried out, and the stable isotope labeled snake venom protein with the purity reaching 95% or above and the labeling efficiency exceeding 91% can be obtained.

To provide a high-affinity monoclonal antibody for a His-tag added to the C terminus of membrane protein.SOLUTION: The present invention provides a monoclonal antibody for a histidine tag (His-tag) or an antigen-binding fragment thereof, wherein the monoclonal antibody recognizes, as an epitope, a His-tag added to the C terminus of a membrane protein.SELECTED DRAWING: None

The application discloses a histidine tag diversity nucleic acid aptamer and application thereof, and belongs to the technical field of biological processing. The nucleotide sequence of the histidine tag diversity nucleic acid aptamer is shown in SEQ ID NO. 3, or SEQ ID NO. 4, or SEQ ID NO. 5, or SEQ ID NO. 6. The application also discloses an application of the histidine tag diversity nucleic acid aptamer in preparation of a biosensor element for targeted recognition of a histidine tag-containing protein or polypeptide. The application also discloses an application of the histidine tag diversity nucleic acid aptamer in qualitative analysis, quantitative detection and separation and purification of the histidine tag-containing protein or polypeptide. The histidine tag diversity nucleic acid aptamer is obtained by taking HisA1-T63 aptamer as an initial chain, preliminarily improving the aptamer through sequence truncation, and repositioning a functional module to a loop region for diversity modification, thereby expanding the structural diversity of the histidine tag nucleic acid aptamer.

The invention relates to the technical field of biology, and discloses a recombinant N-acetylglucosamine-1-phosphate uridine transferase (GlmU) mutant and a preparation method thereof. The sequence of the recombinase is formed by connecting an N-acetylglucosamine-1-uridine phosphate transferase (GlmU) mutant (L113V-V172I) from Azotobacter chroococcum, an N-terminal histidine tag (His-tag) and a small ubiquitin-like modified protein tag (SUMO-tag) in series. The invention further discloses a preparation method of the recombinase. Compared with a recombinant wild type N-acetylglucosamine-1-uridine phosphate transferase with the same source, the recombinant N-acetylglucosamine-1-uridine phosphate transferase mutant has higher activity which is 1.5 times of that of the recombinant wild type and higher stability, and the residual activity of the mutant is 66.5% after the mutant is placed at room temperature for 5 days, so that the mutant has a good application prospect in the field of N-acetylglucosamine-1-uridine phosphate transferase. And the soluble expression quantity of the recombinant mutant N-acetylglucosamine-1-uridine phosphate transferase is increased by 5 times.

The invention pertains to His tagged FimHLD R60P protein for use as a medicament, in particular as an antigen in a vaccine to protect against an infection with E. coli.

The invention belongs to the field of biological medicine, and particularly relates to a protein purification method based on an anti-His tag monoclonal antibody. The antibody can recognize histidine tags with high specificity, an affinity chromatography medium prepared with the antibody as a ligand can efficiently purify His tag protein, the problem that metal ions fall off in traditional metal chelating chromatography can be effectively solved, and non-specific adsorption of impure protein is remarkably reduced. The purification method based on the medium has strong targeting capture capability, can greatly improve the final purity and yield of the protein with the histidine tag, and provides a reliable solution for obtaining the high-purity protein.

The invention discloses an HlyA secretion system for increasing the secretion amount of substrate protein and a construction method and application thereof.The HlyA secretion system comprises a first component, a second component and a third component, the first component comprises an inner membrane transporter HlyB and a membrane interstitial adaptor protein HlyD, and the inner membrane transporter HlyB and the membrane interstitial adaptor protein HlyD are obtained by expressing modified hlyBD operons; according to the modified hlyBD operon, a nucleotide sequence for coding a hexa-polyhistidine tag is inserted behind an hlyD gene initiation codon in the hlyBD operon; and a second component: an outer membrane channel protein TolC derived from a host cell. According to the invention, the HlyD protein is specifically modified, so that the secretion amount of the HlyA secretion system to the substrate protein is obviously increased.

This invention discloses a metal ion-driven meliosteum peptide nanoassembly, its preparation method, and its applications, relating to the field of agricultural biotechnology. The meliosteum peptide nanoassembly is formed by the self-assembly of a hexahistidine-tagged meliosteum peptide with divalent metal ions through coordination. The average particle size of the meliosteum peptide nanoassembly is 20 nm–30 nm. The meliosteum peptide nanoassembly obtained by this invention through the self-assembly of a hexahistidine-tagged meliosteum peptide with zinc ions exhibits excellent antibacterial, anti-biofilm activity, stability, and biocompatibility, and can be used to control plant bacterial diseases caused by Xanthomonas and other fungi.

The present application relates to the technical field of biology, and particularly relates to a method for stably improving horseradish peroxidase-labeled streptavidin titer. The method provided by the present application is to design streptavidin with different histidine tag lengths, and then couple with sodium periodate-oxidized horseradish peroxidase, so that high-titer horseradish peroxidase-labeled streptavidin can be obtained. The present application also discloses application of the enzyme-labeled conjugate in a magnetic microsphere platform streptavidin-biotin system. Experimental results prove that the method provided by the present application can stably prepare high-titer and low-background enzyme-labeled conjugate, and the enzyme-labeled conjugate can be widely applied in the streptavidin-biotin system.

The invention relates to a p-toluenesulfonyl functionalized fluorescent probe capable of being specifically combined with a histidine label and application of the p-toluenesulfonyl functionalized fluorescent probe in single-molecule FRET (Fluorescence Resonance Energy Transfer) research. The probe is based on a Cyanine 5 dye, and can realize covalent binding with a protein with a His-tag by performing chemical modification on toluenesulfonyl. Compared with a traditional Ni < 2 + >-NTA non-covalent fluorescent probe, the probe provides higher binding stability and site specificity, and is suitable for high-resolution single-molecule fluorescence imaging of protein. The invention further establishes a protein fluorescence labeling strategy which is suitable for smFRET analysis and is universal, simple and convenient to operate and stable in labeling, and a novel technical means is provided for exploring protein conformation change and a dynamic regulation and control mechanism.

The application discloses a method for expressing BirA enzyme by using prokaryotic Escherichia coli and purifying the BirA enzyme, and relates to the technical fields of genetic engineering and protein engineering. A recombinant expression vector for expressing the BirA enzyme is a pET-28A-BirA plasmid which is constructed by inserting a BirA enzyme coding gene into a multiple cloning site of a pET-28A vector. An 8× histidine tag coding nucleotide sequence is fused to a C terminal of the BirA enzyme coding gene. The nucleotide sequence of the BirA enzyme coding gene is shown as SEQ ID NO:1. The prokaryotic expression system is simple in culture condition, short in cycle, and low in cost, and is suitable for large-scale preparation. The 8× histidine tag is introduced into the C terminal of the BirA enzyme gene, so that the expression product can be purified by one-step nickel column affinity chromatography, and the purification process is simple and high in efficiency.

The present invention relates to a method for isolating and purifying a protein with at least two histidine tags, in particular to a method for isolating and purifying a protein with at least two histidine tags in solution by using cations.

Disclosed is using a TEV protease (wild type or mutant) suited for removal from a reaction mixture, wherein the TEV protease displays a dual affinity tag including a chitin binding domain (CBD) tag and a histidine tag, preferably where the CBD tag precedes the N-terminus of the protease and the CBD tag is preceded by the histidine tag, which is preferably a 6-mer histidine tag; and optionally further including linkers, preferably Gly-Ser linkers, and more preferably a 6-mer Gly-Ser linker, between the tags. A linker, also preferably a Gly-Ser linker, can also follow the CBD tag and precede the protease portion. In certain embodiments, no such linkers are present and in other embodiments, only one such linker is present.

The invention belongs to but is not limited to the technical field of gene engineering, and discloses an anguilla japonica Activin B recombinant protein and an expression method and application thereof, the anguilla japonica Activin B recombinant protein comprises an anguilla japonica Activin B mature peptide amino acid sequence, a 6 * histidine tag at the N end and SUMO dissolution promoting polypeptide; the amino acid sequence of the anguilla japonica Activin B recombinant protein is SEQ ID NO: 1, and the amino acid sequence of the anguilla japonica Activin B mature peptide is SEQ ID NO: 2. The prokaryotic expression vector pET-His-SUMO-Activin B of the Japanese eel Activin B is constructed for the first time, the Activin B recombinant protein with the amino acid sequence consistent with the mature peptide of the Japanese eel Activin B is obtained, and the Activin B recombinant protein can be applied to ovarian development regulation and artificial propagation research of the Japanese eel.

The application discloses a novel coordination peptide QEY127 and application thereof in assembling a luminescent material with a rare earth ion. The coordination peptide QEY127 is composed of one start codon coding methionine (M), one 6-polyhistidine tag (6X His) and 10 QSEPGDPGEPSY (QEY12), and has a length of 127 amino acid residues; the coordination peptide QEY127 gene is obtained through a whole gene synthesis method, and the codon composition is optimized. The yield of the coordination peptide QEY127 is 100-1200 mg / L of fermentation liquor, and the purity is 82-95%. The coordination peptide QEY127 has the characteristics of glutamine transaminase (TG enzyme) catalyzed crosslinking, high elasticity and formation of a luminescent coordination compound with a rare earth metal, and has a wide application prospect in the fields of biological medical treatment and luminescent materials.

Disclosed herein are compositions and constructs related to a polypeptide, PT-X'n-PTA-Hm (1 ), or pharmaceutically acceptable salts thereof, wherein PT is a peptide transport or fragment thereof; X'n is linker comprising a triazole moiety, n = 0 or 1; PTA is a peptide therapeutic agent selected from the group consisting of a modified semaglutide, a modified tirzepatide, caplacizumab, ozoralizumab; and Hm is a histidine tag where m is 0 to 6; as well as uses thereof.

The invention provides an antibody SEC55, a fusion protein SEC55-vHRP, a preparation method and an application. The fusion protein comprises an IgG antibody signal peptide, a hemagglutinin tag, the antibody SEC55, a connecting peptide, horse radish peroxidase and a histidine tag. The fusion protein can be used for detecting staphylococcus aureus enterotoxin C in food. The fusion protein SEC55-vHRP is provided with a horse radish peroxidase label, so that secondary antibody incubation is not needed, subsequent chromogenic reaction can be directly carried out after primary incubation is finished, the detection time is shortened, and the fusion protein SEC55-vHRP has a wide application prospect.

Either wild type or mutant Serratia Marcescens Nuclease (“SMN”) is engineered to display a C-terminal Chitin Binding Domain (CBD-tag), followed, at the C-terminus side of the CBD-tag, by a poly-histidine tag (His-tag), where the His-tag is preferably a 6-mer and preferably is preceded by a Gly-Ser linker, thereby generating a recombinant SMN protein that retains dual affinity tags (a CBD-tag and a His-tag) at the C-terminus, to make it easily removed from a reaction solution following digestion of nucleic acids in the reaction mixture. It can also be used for binding SMN to a solid support for use in nucleic acid digestion in a sample contacted with the solid support.

This article discloses a material composition and a method for using the composition to treat... For example Methods for treating cancer or inflammatory joint diseases. The composition is a soluble recombinant human flatfoot protein receptor (srhPDPN) comprising a first amino acid sequence at least 95% homologous to SEQ ID NO:1. The srhPDPN may comprise a tag peptide or polypeptide (such as a multihistidine tag) fused to the first amino acid sequence. The srhPDPN may comprise a signal peptide fused to the first amino acid sequence. The signal peptide may comprise an amino acid sequence at least 95% homologous to SEQ ID NO:2. Multiple threonine residues of the srhPDPN may be partially glycosylated with sialic acid, and in some embodiments, 17 threonine residues of the srhPDPN are partially glycosylated with sialic acid. Treatment may include allowing the composition to act as a decoy receptor or ligand trap to inhibit PDPN signaling.

Provided are an enzyme immobilization carrier and a preparation method therefor, an immobilized enzyme and a preparation method therefor, and use thereof. The enzyme immobilization carrier comprises a resin matrix and a divinyl sulfone group modified on the resin matrix by means of a covalent bond. In order to realize the directed immobilization of an enzyme, the divinyl sulfone group is loaded on the resin matrix by means of the covalent bond. In a subsequent enzyme immobilization reaction, by using specific binding of the divinyl sulfone group to a histidine tag, the divinyl sulfone group in the enzyme immobilization carrier is connected to a specific amino acid residue in an enzyme molecule by means of a covalent bond, without reacting with an active group in the enzyme molecule, thereby effectively avoiding excessive influence of the enzyme immobilization carrier on enzyme conformation, and thus retaining a high catalytic activity of the enzyme. Compared with a traditional metal chelating type resin-enzyme molecule binding mode, the present invention shows better immobility.

The invention discloses a method for separating DHA and EPA in fatty acid or separating DHA-EE and DPA-EE in fatty acid ethyl ester. DHA is separated from a DHA and EPA mixed solution through affinity difference between FABP protein and fatty acid, and the problems of high energy consumption and easy oxidation of a traditional separation method are avoided. The preparation method comprises the following steps: firstly, respectively adding strep-tag II and six histidine tags to N ends and C ends of FABP3 and FABP7 protein sequences, inserting the FABP3 and FABP7 protein sequences into a plasmid pET-28a (+), and expressing FABP3 / FABP7 protein with tags through escherichia coli; then, the affinity (Kd) of FABP3 and FABP7 with DHA / EPA is measured through a BLI experiment, and the affinity (Kd) is expanded to the affinity (Kd) of DHA-EE / DPA-EE; finally, the protein is coupled to a nickel column, and DHA / EPA is subjected to quantitative analysis through GC-MS to evaluate the protein separation effect.

The invention discloses a histidine molecular imprinting monolithic column as well as a preparation method and application thereof, and belongs to the technical field of high polymer materials. Polysaccharide is dissolved in water, the physical state of the polysaccharide is changed, and a polysaccharide aqueous solution is obtained; the preparation method comprises the following steps: reacting a cross-linking agent with a polysaccharide aqueous solution under alkaline conditions to obtain an epoxy group-containing polysaccharide aqueous solution, cross-linking the epoxy group-containing polysaccharide aqueous solution with a histidine molecular imprinting template, cooling and curing to obtain the histidine tag template-containing molecular imprinting monolithic column, and separating the histidine tag template-containing molecular imprinting monolithic column from the histidine tag template-containing molecular imprinting monolithic column to obtain the histidine tag template-containing molecular imprinting monolithic column. And finally, cleaning the histidine tag molecular imprinting template to obtain the histidine molecular imprinting monolithic column. The whole preparation process is simple and is completed through continuous reaction under the condition of an aqueous solution, no heavy metal such as Cu or Ni and organic solvents participate in production and are not left, and the preparation method has the advantages of being safer, environmentally friendly and economical. Therefore, the method has a good application prospect.

The application provides an analysis method for determining histidine tag protein concentration based on IMAC-HPLC and application thereof, and the analysis method comprises the following steps: (1) preparing reference solution, quality control solution and sample solution to be detected, and performing IMAC-HPLC detection; (2) taking the sample amount of the reference solution as the abscissa X and the peak area of the histidine tag protein as the ordinate Y, fitting a linear equation to obtain a standard curve; (3) bringing the peak area of the histidine tag protein in the quality control solution into the standard curve to calculate the measured sample amount of the histidine tag protein in the quality control solution, and the ratio of the measured sample amount to the theoretical sample amount is the recovery rate; and bringing the peak area of the histidine tag protein in the sample solution to be detected into the standard curve to calculate the measured sample amount of the histidine tag protein in the sample to be detected, and the ratio of the measured sample amount to the sample injection volume is the histidine tag protein concentration.

Quantum dots (QDs) and nanoplatelets (NPLs) are two types of nanoparticles used as scaffolds for enzymes operating in enzymatic cascades. Combinations of QDs and NPLs were surprisingly found to operate synergistically to create a greater enhancement than either alone when operating as scaffolds for enzymatic cascade reactions. A process involves providing an enzymatic cascade including a cluster of nanoparticles including both QDs and NPLs and having a plurality of enzymes bound thereto, the enzymes configured as an enzymatic cascade, such that the product of a first enzyme is a substrate of a second enzyme; contacting the cascade cluster with a substrate of the first enzyme; and allowing a reaction to proceed so that each of the plurality of enzymes acts in succession to produce an end product. The enzymes are bound to the nanoparticles via metal affinity coordination between histidine tags on the enzymes and zinc-containing surfaces of the nanoparticles.