6 results about "Cytoplasmic protein" patented technology

The invention discloses an engineered protein and application thereof in mediating non-ubiquitination degradation, and belongs to the technical field of biological medicine. The structural domains CATCH1 and CATCH2 of the MIDN protein and the region between the structural domains CATCH1 and CATCH2 are modified for the first time, the modification mode comprises the steps that the structural domains CATCH1 and CATCH2 and the region between the structural domains CATCH1 and CATCH2 are replaced with antibodies or polypeptides, then structure or sequence optimization is conducted on the basis, and the obtained MIDN engineered protein not only retains the proteasome binding capacity of MIDN, but also has the advantages of being capable of improving the protein quality and the like. According to the present invention, the MIDN-based target protein degradation system has the MIDN-based target protein degradation system, can expand the target range to the membrane protein or the cytoplasm protein, can improve the degradation efficiency of the MIDN-based target protein degradation system, further has the antibody-mediated target protein specificity so as to achieve the pathological / physiological protein distinguishing effect, and does not have the obvious toxicity; therefore, the engineered protein provided by the invention has a good application prospect.

The invention discloses a method for enriching and extracting soluble protein in a tissue sample, which comprises the following steps: adding the tissue sample into a low-salt buffer solution, homogenizing, centrifuging, collecting supernatant precipitate, resuspending the precipitate by using Triton, and obtaining the supernatant to obtain an extracting solution; adding anhydrous acetonitrile to precipitate protein and collecting precipitate; carrying out proteolysis reductive alkylation and quenching reaction treatment on the precipitate to obtain a peptide fragment sample; desalting by using a peptide fragment sample desalting cleaning solution; after washing, adding a desalted eluent for eluting, collecting an eluent containing the peptide fragment, and drying to obtain a desalted peptide fragment; performing reversed-phase fractionation chromatography under an alkaline condition; and finally, LC-MS / MS detection is carried out. Under the condition that a strong denaturing agent is not introduced, secretory protein, cytoplasm protein and membrane protein can be selectively enriched, meanwhile, most of cell nucleus protein and tissue interstitial fibrin are effectively removed, and therefore the detection capacity of follow-up mass spectrometric detection on extremely-low-abundance circulating markers is remarkably improved.

The invention relates to application of a nano-micelle based on functional nucleic acid in targeted protein degradation. Specifically, the invention provides a nucleic acid aptamer-connexon-lipid nanostructure, constructs editable universal platforms respectively utilizing two degradation systems of lysosome and ubiquitin-proteasome, and is respectively used for degradation of membrane protein and cytoplasm protein. A new thought is provided for avoiding the problems of degradation system overload and safety possibly caused by a single degradation path in traditional targeted protein degradation, and technical support is provided for treatment of diseases such as tumors.

Compositions and methods are provided for the cell-mediated targeted killing of diseased cells based on the presence of an intracellular antigen, rather than a surface-bound marker. The targeting cells are modified to express a cytotoxic protein that is delivered into a targeted cell, and after delivery is selectively activated by the presence of a cytoplasmic protein of interest. In one embodiment of the invention, the cytotoxic molecule is a Granzyme B (GrB) polypeptide. In the compositions of the invention, GrB is modified to render its cytotoxic enzymatic functions inactive, until the presence of an intracellular antigen unlocks the GrB molecule to enable enzymatic activities.

Provided are a nanocomposite for targeted degradation of a pathogenic protein, a preparation method therefor, and use thereof, which pertain to the technical field of nanobiological drugs. The nanocomposite for targeted degradation of the pathogenic protein is provided, which comprises a nanocarrier and a protein-targeting binding peptide grafted on the nanocarrier. The nanocarrier is a nanoassembly of maleimide-polyethylene glycol-polylactic acid and cationic lipids; on the other hand, the use of the nanocomposite in the preparation of drugs including an anti-tumor nanodrug and a Huntington's disease inhibiting drug is provided. The nanocomposite can simulate a key receptor protein in a selective autophagy pathway, so that the pathogenic protein to be degraded can be brought into an autophagosome to be degraded by means of an autophagy pathway, thereby effectively solving the problem that PROTACs cannot degrade large-molecular-weight protein aggregates and LYTACs cannot degrade cytoplasmic proteins.

The invention discloses a low-abundance cell nucleus enrichment method which comprises the following steps: (1) splitting a tissue sample to obtain a cell nucleus with extracellular cytoplasm retained; and (2) marking and sorting the cell nucleuses obtained in the step (1) to obtain enriched low-abundance cell nucleuses. According to the method, a mild dissociation technology is used, so that more cytoplasm components can be reserved in cell nucleuses, and then the cell nucleuses are used for marking cytoplasm proteins and are combined with flow sorting, so that directional enrichment of the cell nucleuses with low abundance and specific functional states can be effectively realized. Through the whole process optimization, the method can ensure the completeness of the cell nucleus to the maximum extent, takes downstream sequencing application into consideration, and can obtain high-quality mononuclear transcriptome sequencing data by combining with the screening of cytoplasm sequencing data. Moreover, according to the method, directional enrichment is realized through the cytoplasmic protein, the use cost can be effectively reduced, and the method is suitable for large-scale clinical sample data acquisition and has higher flux compared with a single space transcriptome slice.