3 results about "Histidine residue" patented technology

The invention relates to the field of protein engineering, in particular to a T cell receptor engineering transformation method and application thereof. The T cell receptor engineering transformation method provided by the invention comprises the following steps: obtaining a CDR region of a given T cell receptor sequence through a database, mutating one or more amino acid residues in the CDR region into histidine, and establishing a first T cell receptor mutation library; the T cell receptor engineering transformation method is based on a histidine scanning method, TCR engineering transformation independent of a three-dimensional structure is achieved, the defect of high affinity is overcome, low-affinity and high-activation TCR transformation is achieved, and more choices are provided for clinical application.

This invention relates to the field of biopharmaceutical manufacturing technology and discloses a specific antibody against HER2 and its application, comprising: a heavy chain and a light chain variable region, wherein the complementarity-determining region contains a pH-sensitive binding region composed of histidine residues and hydrophobic residue clusters, and the hydrophobic residue clusters are located in the vicinity of the imidazole ring side chain of the histidine residues, in order to establish a desolvation shielding microenvironment with a low dielectric constant. This invention utilizes the physical shielding of the microenvironment against the solvation interference of water molecules in the blood to maintain the electroneutrality of histidine residues in the physiological environment to stabilize the binding affinity, and in the acidic environment, responds to the proton flow to generate charge flipping and hydration volume expansion stress, synergistically driving the antibody to dissociate from the receptor, thereby solving the drug resistance problem caused by target circulation escape, and improving the efficiency of intracellular receptor downregulation while ensuring serum stability.

The subject of this invention is a construct of an expression vector for expressing recombinant µ-conotoxin TIIIA or TIIIAlaMut, characterized in that it comprises the nucleotide sequence of µ-conotoxin TIIIA SEQ ID NO:1 or the nucleotide sequence of µ-conotoxin TIIIAlaMut SEQ ID NO:4, both sequences containing a sequence encoding six histidine residues (6His) at the 5' end, which is linked via a serine-glycine-serine linker (SGS) to a construct encoding a TRX::TIIIA fusion protein or a TRX::TIIIAlaMut fusion protein, wherein the TRX::TIIIA fusion protein comprises the µ-conotoxin TIIIA gene and a gene encoding a leader protein, and the TRX::TIIIAlaMut fusion protein comprises the µ-conotoxin TIIIAlaMut gene and a leader protein, wherein the leader protein is a thioredoxin (TRX) modified by site-directed mutagenesis, wherein the amino acid methionine at position 37 is replaced by lysine. Another subject of the invention is an expression vector comprising a construct according to the invention under the control of a constitutive promoter. Another subject of the invention is isolated *E. coli* cells comprising an expression vector according to the invention. Another subject of the invention is a method for producing µ-conotoxin TIIIA or TIIIAlaMut in *E. coli* using an expression vector comprising a construct according to the invention, characterized in that the method comprises the steps of: a) transforming *E. coli* cells with an expression vector comprising a construct according to the invention under the control of a constitutive promoter, said expression vector encoding a TRX::TIIIA fusion protein having the amino acid sequence SEQ ID NO:2 or having SEQ ID NO:2. a) TRX::TIIIAlaMut fusion protein NO:5; b) Expression of the TRX::TIIIA or TRX::TIIIAlaMut fusion protein; c) Isolation and purification of the TRX::TIIIA or TRX::TIIIAlaMut fusion protein; d) Formation of disulfide bonds by glutathione treatment of the purified TRX::TIIIA or TRX::TIIIAlaMut fusion protein in GSH / GSSG and dialyzing in buffer; e) Cleavage of the TRX::TIIIA or TRX::TIIIAlaMut fusion protein with the formed disulfide bonds by cyanogen bromide; f) Purification of the cleaved TIIIA or TIIIAlaMut peptide.