129 results about "Amino acid replacement" patented technology

This invention provides a genetically modified marine luciferase such as Gaussia luciferase, which has high bioluminescence intensity, and has high bioluminescence stability and/or red-shifted wavelength. Specifically disclosed is a luciferase variant with improved optical property obtained by replacing at least one amino acid residue among the amino acid sequence of a marine luciferase at positions corresponding to positions 89 to 118 in the amino acid sequence of Gaussia luciferase (GLuc), wherein an amino acid residue at a position corresponding to at least one selected from positions 89, 90, 95, 97, 100, 108, 112, 115, and 118 in the amino acid sequence of GLuc is replaced by way of conservative amino acid replacement. The above-mentioned replacement in a marine luciferase improves enzymatic activity of the luciferase. Also disclosed is a bioluminescent probe having an improved optical property, which is produced using the luciferase variant of the present invention.

The invention provides a novel high-affinity completely humanized antibody which can specifically bind with CD26 as well as a preparation method and application thereof and belongs to the technical field of genetic engineering antibodies. The CD26 is a ubiquitous multifunctional II type transmembrane glycoprotein, has various biological functions and can be interacted with various proteins such as ADA, CD45, FAP-alpha and the like. The invention provides the humanized antibody or a fragment thereof, wherein the antibody or the fragment thereof is capable of specifically binding with the human CD26, preferably the CD26 extracellular domain; the amino acid sequence of the antibody or the fragment thereof comprises an amino acid sequence containing a monoclonal antibody or a fragment thereof or a conjugate of the fragment in any one of 6 complementary determining regions in one of SED ID NO: 2, SED ID NO: 3, SED ID NO: 4, SED ID NO: 6, SED ID NO: 7, and SED ID NO: 8, or an amino acid sequence obtained through amino acid replacement or modification. The obtained anti-CD26 single-chain antibody provided by the invention can highly specifically bind with the CD26 and is simultaneously capable of obviously inhibiting the proliferation, the invasion and the metastasis of tumor cells.

The invention relates to engineering organophosphorus hydrolase, nucleic acid, a mutant and an application of the engineering organophosphorus hydrolase and the mutant and belongs to the technical field of bioengineering. The engineering organophosphorus hydrolase is an organophosphorus hydrolase mutant constituted by new amino acid sequences formed after amino acid substitution of one or more of amino acid residues on the 55th site, the 57th site, the 83th site, the 114th site, the 137th site, the 188th site or the 262th site of an amino acid sequence shown as SEQ ID No.1, and the activity of the engineering organophosphorus hydrolase is improved. Compared with the prior art, the organophosphorus hydrolase mutant has the advantages that the malathion catalytic activity is improved by 36.7 times, the mutant keeps excellent thermal stability of a female parent, and the half-life period at 50 DEG C reaches 65 h. The organophosphorus hydrolase mutant can efficiently degrade common organophosphorus pesticides such as malathion and the like, has excellent thermal stability and has very good application prospect in the field of degradation of the organophosphorus pesticides.

The invention provides a novel full-humanized antibody which is high in affinity and can be specifically combined with CD26, and a preparation method and application thereof, and belongs to the technical field of a genetically engineered antibody. CD26 is a ubiquitous multifunctional II-type transmembrane glycoprotein, has a plurality of biological functions, and can interact with a plurality of proteins, such as ADA, CD45, FAP-alpha and the like. The invention provides an antibody from a human source or a segment thereof. The antibody or the segment thereof is specifically combined with human CD26, and preferably specifically combined with a CD26 extracellular region; an amino acid sequence of the antibody or the segment thereof comprises a monoclonal antibody which is selected from any region of six complementary determining regions containing a group of sequences of SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:7 and SEQ ID NO:8, or a segment thereof or a conjugate of the segment thereof, or an amino acid sequence which is obtained by amino acid replacement or modification. The anti-CD26 single-chain antibody obtained by the method is highly specifically combined with the CD26, and meanwhile, multiplication of tumor cells can also be obviously inhibited.

The invention provides an engineered Cas12i nuclease, which comprises one, two, three or four of following mutations based on reference Cas12i nuclease: (1) replacing one or more amino acids which interact with PAM in the reference Cas12i nuclease with positively charged amino acids; and/or (2) replacing one or more amino acids which participat in opening DNA double chains in the reference Cas12i nuclease with amino acids with aromatic rings; and/or (3) replacing one or more amino acids which are located in the RuvC structural domain and interact with the single-stranded DNA substrate in the reference Cas12i nuclease with positively charged amino acids; and/or (4) replacing one or more amino acids which interact with the DNA-RNA double helix in the reference Cas12i nuclease with positively charged amino acids. Particularly, the reference Cas12i nuclease is natural Cas12i nuclease, such as natural Cas12i2 nuclease, and the amino acid sequence of the reference Cas12i nuclease is as defined in SEQ ID NO. 1.

The invention provides a DNA polymerase with DNA or RNA as a template. The DNA polymerase is obtained through amino acid replacement, including G198W, V222I, E306K, Q354E, A381E and E582K, of klenow fragments of escherichia coli polymerases I. The invention also provides a primer with a stem loop structure and used for constant-temperature nucleic acid amplification. The invention further providesa nucleic acid test technique and a test kit both based on combination between rapid constant-temperature nucleic acid amplification and a Cas test system. The DNA polymerase, the nucleic acid test technique and the test kit can be used for testing nucleic acid targets in test samples at a constant temperature, have the advantages of low cost, short test time, simplicity and convenience in operation, high specificity, high sensitivity and the like, and are particularly applied to POCT (point-of-care testing).

A method for predicting an amino acid sequence compatible with a three-dimensional (3D) structure of a protein. A reduced virtual representation of the 3D structure is constructed, and, for each position along the representation, its solvent accessibility is determined. For each position along the structure, an amino acid residue is randomly selected from a predefined group of amino acids having a solvent accessibility compatible with the solvent accessibility of the position. A Monte-Carlo simulation is performed on this devised protein in which an amino acid at a particular position is sequentially replaced with other amino acids having the same solvent accessibility, and an energy score is calculated for each rotamer. The lowest scoring rotamer for this position is then selected The Monte-Carlo simulation is repeated for each position along the sequence, to obtain an amino acid sequence with the lowest total energy score.

Belonging to the field of biomedicine, the invention provides an endomorphin analogue with the first position, the second position and fourth position respectively modified by tyrosine or 2, 6-dimethyltyrosine, N-methyl-D alanine and alpha-alkenyl-beta-amino acid and a synthesis method thereof. The endomorphin analogue is obtained by substituting the first position amino acid of endomorphin (EM1 and EM2) with tyrosine or 2, 6-dimethyltyrosine, substituting the second position amino acid with N-methyl-D alanine, and substituting the fourth position amino acid respectively with phenyl or 2-furyl replaced alpha-alkenyl-beta-amino acid, and is named as MEL-N16 series. Radioligand receptor binding experiment, isolated organ bioassay, in-vitro enzymolysis stability and warm bath tail flick analgesic experiment results show that the endomorphin analogue synthesized by the method provided by the invention has higher affinity than an endomorphine matrix, high enzymolysis stability and high analgesic activity, and has very good clinical application value in preparation of analgesic drugs.

The invention provides endomorphin-1 modified by unnatural alpha-alkenyl-beta-amino acid, which is characterized in that fourth amino acid of the endomorphin-1 is respectively formed by replacing the alpha-alkenyl-beta-amino acid replaced by phenyl, 2-furan, 3-chlorphenyl, 1, 3-dioxophenyl. Pharmacological activity identity is conducted on the endomorphin-1 modified by the unnatural alpha-alkenyl-beta-amino acid by radiation ligand receptor combination experiments, isolated organ biological assays, cAMP accumulation experiments, separation enzymolysis stability and analgesic tests with water baths. As the results show, the novel analogue has the advantages of high affinity, high enzymolysis stability and high analgesic activity compared with endomorphin-1, thereby having high application value in preparation of polypeptide analgesic medicine.

According to the invention, mutation of several sites of Ser358Asp, Thr480Asn, Asp533Glu and Ala539Gly is carried out on a wild type Bst DNA polymerase sequence, then 292-305 amino acid EGLLKVVRPDTKKV of the Bst DNA polymerase subjected to point mutation is replaced with DPLPDLIHPRTLRL, a DNA binding protein is fused at the C end of the mutated Bst DNA polymerase sequence, an HP47 polypeptide sequence (SEQ ID No.17) is fused at the N end of the mutated Bst DNA polymerase sequence, a CL7-SUMO-Tag is fused in front of the HP47 polypeptide sequence, and the recombinant mutant Super-Bst (SEQ ID No.16) of the Bst DNA polymerase with high activity and thermal stability is obtained. The thermal stability, specificity, strand displacement capacity, extension capacity and reverse transcriptase activity of Super-Bst are remarkably improved, and Super-Bst can tolerate high salt and various inhibitors and can be massively obtained through prokaryotic expression and affinity purification. The invention also discloses coding DNA of the gene and an ultrafast magnetic bead LAMP detection method.

The invention provides a polypeptide and an application thereof to preparation of medicament for treatment and/or prevention of tumor. An amino acid sequence of the polypeptide is shown in a sequence table SEQ ID No.1, or shown in the sequence table SEQ ID No.1 amino acid sequence with two or more amino acids substituted by unnatural amino acids with connectable side chains, and a derivative comprises a chimeric peptide formed by connection of the polypeptide and a cell-penetrating peptide, a fusogenic peptide formed by the polypeptide and a virus, a methylated polypeptide, a glycosylated polypeptide, and a pegylated polypeptide. The polypeptide or the polypeptide derivative can targetedly increase number of PML nucleosomes, and can be applied to preparation of medicament for treatment and/or prevention of tumor.

Provides an artificial enzyme obtained by improving upon a sequence of a natural nicotine dehydrogenase, wherein the improvement comprises replacing at least one amino acid hindering product release with an amino acid with smaller side chains, thereby improving a catalytic rate.

The invention belongs to the technical field of biological pharmacy, and particularly relates to application of active peptide and a mesenchymal stem cell exosome for improving skin physiological characteristics in drugs or cosmetics. Amino acid replacement is performed on the basis of original active peptide, and other conventional amino acids and unusual amino acids are introduced, so that the antioxidant capacity, collagen secretion promoting capacity and cell proliferation promoting capacity are improved. The effect of combined use between the amino acids and the human umbilical cord mesenchymal stem cell exosome is more obvious, skin injuries caused by ultraviolet rays can be effectively prevented and treated, and the repair process after the injuries is promoted.

The invention discloses a polypeptide to specifically promote degradation of EGFR (epidermal growth factor receptor) protein, or its derivatives, and application of the polypeptide in the preparationof tumor drugs. An amino acid sequence of the polypeptide is shown as in sequence table SEQ ID No. 1; alternatively, two or more amino acids in the amino acid sequence shown as in the sequence table SEQ ID No. 1 are replaced with side-chain-connectable non-natural amino acids; the derivatives include chimeric peptides that are formed by connecting the polypeptide with cell penetrating peptides. The polypeptide or its derivatives can promote degradation of EGFR protein and inhibit EGFR signal pathway activity; therefore, the polypeptide or its derivatives are applicable to the preparation of tumor drugs. Drugs prepared herein are suitable for treating the tumors, such as lung cancer, intestinal cancer, pancreatic cancer, breast cancer, liver cancer, and glioma.

The invention provides a high-affinity human antibody capable of being combined with the specificity of CD26, and a preparation method and applications thereof, belonging to the technical field of a genetically engineered antibody. CD26 is of a ubiquitously multifunctional II type transmembrane glycoprotein, has multiple biological functions, can act with various proteins such as ADA, CD45, FAP-alpha and the like. The invention provides an antibody or a segment thereof from a human, the antibody or segment thereof specifically combines the human CD26, preferably, specifically combines the CD26 extracellular region, the amino acid sequence of the antibody or the segment thereof comprises an amino acid sequence obtained by performing amino acid replacement or modification to the monoclonal antibody or segments thereof or conjugates of the segments in any one of six complementary decision regions containing a group of sequences: SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:7 AND SEQ ID NO:8. The anti-CD26 single-chain antibody has high specificity combination with the CD26, and the proliferation and invasion and metastasis of tumor cells can be obviously inhibited.

The present invention concerns a method of genetic modification of a TGB-3 wild type viral sequence for reducing or suppressing the possible deleterious effects of the agronomic properties of a transformed plant or plant cell by said TGB-3 viral sequence, comprising the following successive steps: submitting said sequence to point mutation(s) which allow the substitution of at least one amino-acid into a different amino-acid; selecting genetically modified TGB-3 wild type viral sequences having said point mutation(s) and which are not able to promote cell-to-cell movement of a mutant virus having a dysfunctional TGB-3 wild type viral sequence, when expressed in trans from a replicon; further selecting among said genetically modified TGB-3 viral sequences, the specifically genetically modified sequence which inhibits infection with a co-inoculated wild type virus when the mutant form was expressed from a replicon, and recovering said specifically genetically modified TGB-3 viral sequence. The invention further relates to genetically modified TGB-3 viral sequences suitable to induce gene silencing. In particular hairpin constructs based on such sequences proved highly efficient to induce a PTGS mechanism and degradation of the whole of RNA2 thereby. When plants are transformed accordingly the spread of the virus in the plant is significantly reduced or blocked.