9 results about "Sortase A" patented technology

The application relates to the technical field of biological medicine, in particular to a compound with a conjugated structure containing an alpha, beta-unsaturated aldehyde and application thereof. The compound can covalently modify Sortase A enzyme of propionibacterium acnes through alpha, beta-unsaturated aldehyde, inhibit transpeptidation catalysis and adhesion of gram-positive bacteria (for example, methicillin-resistant staphylococcus aureus, streptococcus mutans and propionibacterium acnes), block anchoring of membrane proteins, inhibit growth of gram-positive bacteria or kill bacteria; conjugated groups with different electric effects and steric effects are screened to adjust the covalent modification efficiency of alpha, beta-unsaturated aldehyde and Sortase A enzyme, strengthen the activity of the compound in inhibiting Sortase A enzyme, thereby improve the antibacterial activity of the compound, and further make the compound with the conjugated structure containing the alpha, beta-unsaturated aldehyde in the application be applied to the fields of biological medicine and cosmetics as an antibacterial component.

A sortase variant with improved catalytic activity and stability, in particular said mutant further comprises the mutations M155V / L200F or M155V / K162P, according to SEQ ID NO: 1, with respect to the parent sortase (SEQ ID NO: 2). Also the use of said sortase mutant is involved.

The invention discloses a preparation method of a proteoglycan protein conjugate carrying a specified connection site. The method comprises the following steps: carrying out recombinant expression on a recombinant tetanus toxin heavy chain C fragment rTTHc containing a Sortase A 7M ligase recognition sequence, introducing O, O '-(ethane-1, 2-diyl) bis (hydroxylamine) as a nucleophilic reagent into the C tail end of the rTTHc under the action of ligase Sortase A 7M, and carrying out specific condensation reaction on the C tail end of the rTTHc as a unique chemical reaction site and polysaccharide carrying an aldehyde group functional group to obtain the tetanus toxin heavy chain C fragment rTTHc. Therefore, polysaccharide protein conjugates with definite polysaccharide and protein connection sites are produced. According to the invention, the problem that the key antigen epitope of the carrier protein and the heterogeneity of the product are possibly damaged by the traditional random modification strategy is solved, the antigen epitope of the carrier protein is prevented from being damaged to a great extent, and a reliable technical scheme is provided for preparing the polysaccharide conjugate vaccine with high quality and high immunogenicity.

The invention relates to the technical field of biology, and discloses a construction method and application of a BCMA full-length receptor in a nano phospholipid plate. The construction method of the BCMA full-length receptor comprises the following steps: reacting a BCMA extracellular domain, a BCMA transmembrane region and an intracellular domain containing site-specific mutagenesis, CaCl2 and sortase A, after the reaction is finished, purifying by using a nickel affinity column, collecting an imidazole elution component, and enriching to obtain the BCMA full-length receptor in the nano phospholipid plate. According to the construction method, the extracellular domain of the BCMA and the BCMA transmembrane region and the intracellular domain in the nano phospholipid plate are connected through Sortase A enzyme, the BCMA full-length receptor is obtained in an in-vitro near-physiological environment, the method is simple, protein is easy to obtain, and the defect that the transmembrane region is poor in solubility is overcome; meanwhile, the structure-function of the BCMA can be studied in vitro by marking the functional structure domain of the BCMA at a fixed point.

This invention discloses a nanobody Nb.TB1, engineered plant exosomes, their preparation methods, and applications, belonging to the field of biomedical technology. The amino acid sequence of the nanobody Nb.TB1 is shown in Seq ID No. 1. Furthermore, this invention proposes a method for preparing engineered plant exosomes, comprising the following steps: S1, extraction of Gastrodia elata exosomes: extracting Gastrodia elata exosomes from Gastrodia elata using an enzymatic method; S2, expression and purification of the blood-brain barrier-crossing nanobody fusion protein with Sortase A ligase; S3, surface modification of the Gastrodia elata exosomes. In addition, this invention also proposes an engineered plant exosome prepared by the above method. Furthermore, this invention proposes the application of the above-mentioned engineered plant exosomes in the preparation of targeted therapeutic drugs for central nervous system diseases. The engineered plant exosomes prepared by this invention exhibit high blood-brain barrier crossing efficiency.

The invention discloses a Sortase A transpeptidase mutant and application thereof in the technical field of bioengineering, the Sortase A transpeptidase mutant is obtained through single mutation or double mutation on the basis of a protein sequence shown in SEQ ID NO.2, the protein sequence shown in SEQ ID NO.2 is a truncated mutant at the 60th-206th site made on wild type Sortase A transpeptidase shown in SEQ ID NO.1, and the Sortase A transpeptidase mutant is obtained through single mutation or double mutation on the basis of the protein sequence shown in SEQ ID NO.2. The Sortase A transpeptidase mutant provided by the invention has higher yield, activity and thermal stability, and has wider application conditions. Besides, a cysteine is introduced to the C terminal of the Sortase A transpeptidase mutant protein, and the Sortase A transpeptidase mutant is fixed on the magnetic beads by utilizing the principle of sulfydryl covalent coupling, so that the application value of the Sortase A transpeptidase mutant in protein labeling is realized.

The invention relates to application of dryopteris crassirhizoctonin ABBA in preparation of a Listeria monocytogenes infection inhibitor, the compound can effectively inhibit activity of key virulence factors of Listeria monocytogenes, namely hemolysin (LLO) and sortase A (SrtA), and reduce hemolytic activity by blocking LLO oligomerization. The formation of a SrtA-mediated biological membrane and the adhesion and invasion of bacteria to host cells are inhibited. On the premise of not influencing the normal growth of bacteria, the ABBA obviously relieves cytotoxicity, inflammatory response and tissue damage caused by the listeria monocytogenes, and improves the survival rate of a host in an infection model. The inhibitor is derived from natural plants, is high in safety, and provides a new way for the development of anti-Listeria monocytogenes drugs.

A compound A-L-B, wherein A is a nanoparticle (NP)-forming unit, preferably a NP-forming polypeptide or a NP-forming protein, preferably a NP-forming polypeptide or a NP-forming protein having its C-terminus covalently bound to the N-terminus of L via a peptide bond; L is a polypeptide having the following amino acid sequence written in the single letter code z1–G X1 X2 G X3 G X4 X5 G X6 G X7 G–z2, wherein X1 = any amino acid except H, C, and W, X2 = any amino acid except H, C, and W, X3 = any amino acid except H, C, and W, X4 = any amino acid except H, C, and W, X5 = any amino acid except H, C, and W, X6 = any amino acid except H, C, and W, X7 = any amino acid except H, C, and W, z1 represents the N-terminus of the polypeptide, or a group consisting of 1 to 10 amino acids, and z2 represents the C-terminus of the polypeptide, or a modification of the C- terminal carboxyl group of the polypeptide, which modification (i) forms together with the carboxyl group of the C-terminal amino acid of the polypeptide a moiety having the structure -C(O)-O-R1 or -C(O)-NR2R3, wherein R1 is a functional group selected from the group consisting of -(CH2)n-N3, -(CH2)n-C≡CH, -(CH2)n-difluorooctyne (DIFO), and -(CH2)n-dibenzylcyclooctyne (DIBO); and wherein one of R2 and R3 is H and the other one is a functional group selected from the group consisting of -(CH2)n-N3, -(CH2)n-C≡CH, -(CH2)n-difluorooctyne (DIFO), and -(CH2)n-dibenzylcyclooctyne (DIBO); wherein n = 1, 2, 3, 4, or 5; or (ii) is a polypeptidic catcher group which is able to form an isopeptide bond with a polypeptidic tag group in a catcher / tag pair reaction; or (iii) is a Staphylococcus aureus sortase A transpeptidase recognition site consisting of the amino acids LPXTG, being attached to the C-terminus of the polypeptide via a peptide bond; wherein the side chain of each amino acid of the polypeptide L independently of each other may be chemically modified, in particular phosphorylated, amidated, acetylated, glycosylated, PEGylated, HESylated or combinations thereof; and B is a compound having biological activity, preferably a protein, which preferably has its N-terminus covalently bound to the C-terminus of L via a peptide bond; wherein the N-terminus of L is covalently bound to A, preferably via a peptide bond, and wherein the C-terminus of L is covalently bound to B, preferably via a peptide bond.

The invention provides a polypeptide compound as well as a method for preparing cyclopeptide oxygen ester and application thereof, and relates to the field of chemoenzymatic modification of a gene encoding peptide library. According to the preparation method of the cyclopeptide oxygen ester, the preparation of the cyclopeptide molecule connected with the side chain to the side chain oxygen ester by a Sortase A enzyme system is realized for the first time, and the cyclopeptide oxygen ester is applied to the oxygen ester bond cyclization modification of a phage display polypeptide library to generate a gene coding cyclopeptide oxygen ester library and is used for finding the cyclopeptide oxygen ester molecule combined with a target protein. The defects in the aspect of preparation of the cyclopeptide oxygen ester library in the prior art are overcome, the preparation method of the cyclopeptide oxygen ester has mild reaction conditions and high reaction efficiency, and the generated gene coding cyclopeptide oxygen ester library can be used for screening functional cyclopeptide oxygen ester. And a powerful platform technology is provided for the fields of drug discovery and biological materials.