11 results about "Peptide substrate" patented technology

The invention discloses an electrochemical urine biopsy system for cathepsin B analysis and a detection method of the electrochemical urine biopsy system. The electrochemical urine biopsy system comprises gold nanoparticles AuNPs, a peptide-peptide nucleic acid PNA series probe and an electrochemical sensor. The peptide-peptide nucleic acid tandem probe is combined with gold nanoparticles through a mercaptan modified sequence to form a spherical nanostructure with the diameter of 15 nm, a polypeptide substrate sequence serves as a specific reaction substrate of cathepsin B, and PNA provides stability and a signal shielding function. The electrochemical sensor is based on a cutting enzyme-assisted DNAwalker mechanism, signal amplification is started through methylene blue MB release, and high-sensitivity detection of the released PNA probe is achieved. The system provided by the invention has the characteristics of high specificity, high sensitivity and rapid response, can realize quantitative analysis of cathepsin B through noninvasive urine biopsy, and provides a new technical means for early diagnosis and treatment monitoring of tumors.

To provide protease-responsive drug delivery hydrogels, uses thereof, and related methods of their production, more particularly, to provide hydrogels which release anti-inflammatory agents upon reaction with inflammation-related proteases.SOLUTION: A drug-loaded protease-responsive hydrogel is provided, comprising: a) a drug covalently bound to a protease-cleavable peptide anchor having a functional moiety; b) a polymer building block comprising a multi-arm polyethylene glycol (PEG) polymer having at least one functional moiety; and c) a bis-functional crosslinker comprising a peptide substrate flanked by spacer sequences containing functional moieties; wherein the drug and the arm of the multi-arm PEG polymer are covalently bound through the functional moiety of the peptide anchor, and a gel is formed by covalent bonding between the functional moiety of the polymer building block and the functional moiety of the bis-functional crosslinker.SELECTED DRAWING: Figure 19

The invention provides an HPLC (High Performance Liquid Chromatography)-based chymosin activity detection method taking polypeptide as a substrate and application, and relates to the technical field of enzyme activity detection. The method comprises the following steps: mixing 1 mg / mL-2. 5 mg / mL of a substrate polypeptide solution and 0.002 U / mL-0. 02 U / mL of a test solution for enzymolysis, centrifuging to take a supernatant, carrying out HPLC (High Performance Liquid Chromatography) quantitative analysis, and calculating the chymosin activity according to the peak area of the product polypeptide, the sequence of the substrate polypeptide is as shown in SEQ ID No. 1. According to the method for determining the activity of the chymosin based on the HPLC and taking the polypeptide as the substrate, the activity of the chymosin in a chymosin-containing product can be quantitatively detected through the specific 8-peptide substrate of the chymosin, and the method which is easy and convenient to obtain raw materials, high in operability, high in accuracy and good in repeatability is developed for quality judgment of a product taking the chymosin as a main component.

Compositions, methods, and kits are provided for assaying calpain-5 activity and inhibition. In particular, novel peptide substrates are provided for detecting calpain-5, measuring calpain-5 activity, and screening for inhibitors of calpain-5 to identify potential therapeutic agents for treating retinal diseases and other diseases associated with calpain-5 hyperactivity. Additionally, novel inhibitors of calpain-5 are also provided.

Disclosed herein are compositions and methods for the generation of peptide macrocycles from peptide substrates for ATP-grasp enzymes. The peptide substrates contain sufficient linkage cites to produce monocyclic, bicyclic, tricyclic, or multicyclic peptides comprising macrocyclic linkages selected from ω-ester linkages, ω-amide linkages, ω-thioester linkages, and combinations thereof.

A method for determining activity of an acyl transferase enzyme, the method comprising: (i) preparing a reaction mixture comprising: (a) an acyl transferase enzyme, (b) a peptide substrate bound to a fluorophore, wherein the substrate is a cysteine-containing oligopeptide of 5-25 amino acids in length, (c) an acyl-CoA, and (d) a detergent comprising micelles, wherein the acyl transferase enzyme mediates acylation on a cysteine of said peptide substrate to result in association of the peptide with micelles of the detergent with resultant increase in fluorescence polarization; and (ii) measuring fluorescent signal of the reaction mixture; wherein an increase in fluorescence polarization of the reaction mixture compared to fluorescence polarization of a control reaction indicates acyl transferase activity of the acyl transferase enzyme. The above assay method may also be used for screening compounds for their ability to act as inhibitors of an acyl transferase enzyme.

The invention discloses a recombinant fusion protein targeting liver cancer and application of the recombinant fusion protein in preparation of drugs for treating liver cancer, and belongs to the technical field of biological medicines. According to the recombinant fusion protein, human LZTR1WFNLL containing five mutations of K70W, N150F, R494N, P520L and D549L is used as a skeleton, liver cancer targeting peptide SP94, MMP substrate peptide and 9R cell-penetrating peptide are sequentially fused at the N end of the skeleton, the protein stability is enhanced through mutation, the problem that recombinant protein is easy to degrade is solved, and large-scale production and long-term storage are supported; the SP94 is specifically combined with a liver cancer cell surface receptor, so that precise positioning is realized; the MMP substrate peptide responds to tumor microenvironment enzyme digestion to release drugs, and the local concentration is improved; the 9R cell-penetrating peptide promotes protein to penetrate through cell membranes, and the intracellular delivery efficiency is enhanced. The obtained protein is high in biocompatibility and low in immunogenicity. The pharmaceutical composition is accurately enriched in tumor tissues through the mechanism, toxic and side effects of normal tissues are reduced, the anti-tumor curative effect is remarkably improved, and a safe and efficient new strategy is provided for liver cancer treatment.

The application discloses a fluorescence hybridization chain reaction biosensor based on the existence of granzyme B and application thereof, and realizes real-time detection of T cell effector molecules by establishing a fluorescence hybridization chain reaction biosensor based on the existence of active granzyme B, and has the characteristics of short amplification time, simple operation, high amplification efficiency, simple reaction system and high economic benefit. By introducing peptide nucleic acid into the hybridization chain reaction system, the cutting of the substrate by granzyme B is successfully used as the starting recognition event of the hybridization chain reaction, and after the cutting of the peptide substrate in the peptide nucleic acid by granzyme B, the foothold is exposed, the strand displacement reaction is mediated, the fluorescence signal is restored after amplification, and a new strategy of using the hybridization chain reaction for detecting active cytokines is provided. The strategy makes the hybridization chain reaction using fluorescence as a signal output mode applicable to active protein detection, and has the advantages of a simple reaction system, few operation steps and low detection cost.

The invention relates to a single-molecule peptide recognition and enzyme activity detection method, in particular to a beta-secretase activity detection method based on fluorescent scintillation fingerprints and deep learning recognition. The method comprises the following steps: marking a spontaneous scintillation type fluorescent dye on a specific peptide substrate, and fixing the specific peptide substrate on the surface of a functionalized slide by utilizing click chemistry to realize stable anchoring of a single peptide molecule; then, a total internal reflection fluorescence microscope is adopted to collect a time sequence fluorescence track, and single molecule scintillation fingerprints before and after enzyme digestion are obtained. And track data is classified and analyzed by combining a deep learning model, so that whether the peptide molecules are subjected to enzyme digestion or not can be accurately judged, and quantitative detection of the enzyme activity is further realized. Compared with a traditional method, the method has the advantage that the detection sensitivity, the specificity and the adaptability to complex samples are remarkably improved. The beta-secretase is used as a model, the verification accuracy rate exceeds 88%, a new technical path is provided for enzyme activity analysis and early diagnosis of related diseases, and the method has a wide application prospect.

Disclosed herein are compositions and methods for the generation of peptide macrocycles from peptide substrates for ATP-grasp enzymes. The peptide substrates contain sufficient linkage cites to produce monocyclic, bicyclic, tricyclic, or multicyclic peptides comprising macrocyclic linkages selected from ω-ester linkages, ω-amide linkages, ω-thioester linkages, and combinations thereof.

This invention discloses a highly active transglutaminase polypeptide substrate and a screening method, belonging to the field of polypeptide screening technology. The screening method is based on molecular docking to obtain five pairs of highly active transglutaminase polypeptide substrates. After enzyme activity determination of the five pairs of polypeptide substrates, the pair with the highest activity was finally determined. The polypeptide substrate includes an acyl donor and an acyl acceptor. The acyl donor sequence is AFQSAY; the acyl acceptor sequence is FMKHKFV. The highly active mTGase polypeptide substrate provided in this application exhibits a 100-fold increased sensitivity to mTG compared to collagen. Using it to test enzyme activity can avoid substrate overabundance caused by non-specific coupling in biological samples, improve the efficiency of enzymatic reactions, and achieve highly sensitive detection of mTGase in the field of food safety.