13 results about "Ligase activity" patented technology

The present invention provides a polypeptide ligase mutant and a method for preparing a polypeptide. The polypeptide ligase mutant comprises: (a) a protein mutated from the amino acid sequence shown in SEQ ID NO: 1, the mutation comprising a mutation occurring at the S307 site; or (b) a protein having 70% or more identity with the amino acid sequence defined in (a) and having polypeptide ligase activity. The present invention can solve the problem in the prior art of poor activity of polypeptide ligases, and is applicable to the field of polypeptide synthesis.

A method for producing γ-Glu-Val-Gly is described, wherein the method includes the steps of cultivating a γ-Glu-Val-Gly-producing bacterium belonging to the family Enterobacteriaceae in a culture medium so that the γ-Glu-Val-Gly accumulates in the culture medium or the cells of the bacterium, or both, and collecting the γ-Glu-Val-Gly from the culture medium or the cells of the bacterium, or both. The bacterium has been modified to overexpress a gene encoding a protein having L-threonine 3-dehydrogenase activity and a gene encoding a protein having 2-amino-3-oxobutanoate coenzyme A ligase activity.

Provided herein are compositions, kits, systems, and methods employing single-stranded end-preserving adaptors. Such single-stranded adaptors are attached to DNA duplex molecules while preserving original 5′ or 3′ single-strand protruding ends (e.g., present in cell-free DNA) by attaching such adapters to 3′ ends the DNA duplex molecules using a single strand ligase that has step 3 ligase activity, but not step 2 adenylyl transfer activity, and attaching such adapters to 5′ ends of the DNA duplex molecules using a ligase enzyme (e.g., a circligase), thereby forming loop-like structures on one or each end of the DNA duplex molecules. In further embodiments, the loop-like structures are cleaved (e.g., by an endonuclease) as the single-stranded adapters have a cleavable portion, thereby generating a two-part adapter on one or both ends of the DNA duplex molecules that preserves the initial 5′ or 3′ single-strand protruding ends, along with any methylation present.

To provide methods for degrading target surface proteins in the ubiquitin pathway using bispecific binding agents or immunoconjugates that bind to the target surface proteins and membrane-bound ubiquitin E3 ligase.SOLUTION: The present invention relates, inter alia, to methods of degrading target surface proteins in the ubiquitin pathway using bispecific binding agents or immunoconjugates that bind to target surface proteins and membrane-bound ubiquitin E3 ligase. The present invention also relates to methods of degrading target surface proteins in the ubiquitin pathway using engineered membrane spanning proteins that bind to target surface proteins and exhibit ubiquitin E3 ligase activity. The present disclosure also provides compositions and methods useful for making bispecific binding agents and engineered transmembrane proteins, immunoconjugates, nucleic acids encoding them, host cells genetically modified with said nucleic acids, and methods of modulating the activity of cells and / or treating various diseases, such as cancer.SELECTED DRAWING: Figure 1

In one aspect, the invention relates to substituted 1-phenyl-3-(piperidin-4-yl)urea analogs, derivatives thereof, and related compounds, which are useful as inhibitors of the DCN1-UBC12 interaction inhibitors of DCN1-mediated cullin-RING ligase activity, methods of making same, pharmaceutical compositions comprising same, methods of treating disorders using the disclosed compounds and compositions, methods of treating disorders associated with a DCN1-UBC12 interaction dysfunction, methods of treating disorders associated with a DCN1-mediated cullin-RING ligase activity dysfunction, methods of male contraception comprising the disclosed compounds and compositions, and kits comprising the disclosed compounds and compositions. This abstract is intended as a scanning tool for purposes of searching in the particular art and is not intended to be limiting of the present invention.

This invention discloses a short peptide and its derivatives that mimic the N-terminus of RhoE, and their pharmaceutical applications in the treatment of myocardial hypertrophy, belonging to the field of biomedical technology. This short peptide precisely mimics the amino acid sequence from position 1 to 20 of RhoE, specifically binding to the WW domain of WWP2 and competitively blocking the interaction between the N-Loop and C-Loop of its HECT domain. This causes the WWP2 conformation to change from a self-inhibited "closed" state to an activated "open" state, activating E3 ligase activity. The short peptide retains its binding ability to the cardiomyocyte membrane cavernin CAV3, allowing it to efficiently enter the cardiomyocyte cytoplasm via the cavernin endocytosis pathway, overcoming the deficiency of traditional drugs in clearing intracellular pathogenic proteins. Peptide derivatives constructed by fusing transmembrane peptides such as TAT ​​and T7 further improve intracellular delivery efficiency. Experiments have demonstrated that this short peptide and its derivatives can significantly promote WWP2 self-ubiquitination, enhance cardiomyocyte autophagic flux, clear intracellular pathogenic proteins, and effectively inhibit cardiomyocyte hypertrophy, making it suitable for the preparation of anti-myocardial hypertrophy drugs.

Disclosed herein are compounds, compositions, and methods for inhibiting HECT E3 ubiquitin ligases. In some embodiments, the compounds are formulated as a pharmaceutical composition and administered to a subject in need thereof. In some embodiments, the subject in need thereof is diagnosed with a neurological disease or a cancer characterized by increased or ectopic HECT E3 ligase activity.

This invention discloses a T4 RNA ligase 2 mutant, its preparation method, and its applications, relating to the fields of biology and enzyme engineering. The T4 ligase 2 mutant provided by this invention has mutations at at least one of the following positions in the wild-type T4Rnl2 sequence: positions 62, 103, 166, 168, 193, 217, 260, 297, 298, 303, 306, 311, 313, and 318. Compared to the wild type, the mutant exhibits significantly improved thermostability and maintains high ligase activity at higher temperatures, such as 45°C and 50°C. In addition to ligating native nucleic acid sequences, the mutant can also ligate modified non-native nucleic acids at high temperatures. The mutant provided by this invention enables high-temperature synthesis of ribonucleic acid, effectively reducing the generation of byproducts due to base mismatches, improving RNA synthesis efficiency, and facilitating large-scale, high-quality RNA production.

The invention discloses a nucleic acid hybridization capture method and a kit, and the method comprises the following steps: constructing a hybridization reaction system, hybridizing the 5 '-phosphorylated probe to the nucleic acid sample wherein the 5'-portion and the 3 '-portion of the probe are complementary to a first region and a second region, which are not adjacent, in the nucleic acid sample, respectively, and a backbone portion connecting the 5'-portion and the 3 '-portion is not complementary to the nucleic acid sample; reacting the hybrid product under the action of nucleic acid polymerase and / or a polymerase active fragment, and nucleic acid ligase and / or a ligase active fragment to obtain a looping probe; and enriching the ring forming probe. Compared with the prior art, the method disclosed by the invention has the advantages of high specificity, high sensitivity, high flexibility, short process, convenience in operation, wide application scene and the like, and has a good application prospect.

The invention discloses a T4RNA ligase 2 mutant and a preparation method and application thereof, and relates to the field of biology and enzyme engineering, the T4RNA ligase 2 mutant provided by the invention has mutation at at least one site of 62th, 103th, 166th, 193th, 260th, 297th, 298th, 303th, 306th, 311th, 313th and 318th sites of a wild type T4Rnl2 sequence, compared with the wild type, the thermal stability of the mutant is obviously improved, and the T4RNA ligase 2 mutant can be used for preparing a T4RNA ligase 2 mutant. High ligase activity can be maintained at high temperatures such as 45 DEG C and 50 DEG C; in addition to being connected with a natural nucleic acid sequence, the mutant can also be connected with a non-natural nucleic acid with modification at high temperature. According to the mutant provided by the invention, ribonucleic acid can be synthesized at high temperature, so that by-products caused by base mismatch are effectively reduced, the RNA synthesis efficiency is improved, and large-scale high-quality RNA production is facilitated.

This invention discloses a T4 RNA ligase 2 mutant, its preparation method, and related applications, relating to the fields of biology and enzyme engineering. The T4 ligase 2 mutant provided by this invention has a mutation at at least one of the following positions in the wild-type T4 Rnl2 sequence: positions 90, 94, 133, 146, 160, 164, 177, 181, 186, 199, 201, 211, 215, 216, 252, 297, 307, and 330. The mutant provided by this invention exhibits significantly improved thermostability, maintaining high ligase activity even at high temperatures. This mutant can ligate not only native nucleic acid sequences but also efficiently ligate modified non-native nucleic acids. By performing RNA synthesis at high temperatures, byproducts caused by base mismatches can be effectively reduced, significantly improving RNA synthesis efficiency and product purity.

The invention discloses a low-temperature stress molecular regulator ATL31 / 6-CPK28, the plant molecular regulator comprises ATL31 and / or ATL6 and CPK28, ATL31 / 6 and a calcium signal component CPK28 in the plant molecular regulator form a dynamic interaction network: E3 ligase activity of ATL31 is activated under a low-temperature condition, E3 ligase is promoted to be combined with CPK28, ubiquitination degradation of the CPK28 is mediated, and the ATL31 / 6 and the CPK28 are activated under a low-temperature condition. Therefore, a negative feedback regulation loop induced by environmental stimulation is constructed. The invention provides important theoretical breakthrough and molecular targets for plant adversity biology research, has obvious scientific research originality, and also has basic, wide and important application prospects.

Provided herein are bifunctional compounds that bind tau protein and / or promote targeted ubiquitination for the degradation of tau protein. In particular, provided are compounds that can bind tau protein, a protein whose aggregation is implicated in a variety of neurodegenerative disease (e.g., tauopathies), and can promote its degradation by recruiting an E3 ubiquitin ligase (e.g., Cereblon), which can ubiquitinate tau protein, marking it for proteasomal degradation. Also provided are radiolabeled forms of the bifunctional compounds, pharmaceutical compositions comprising the bifunctional compounds, methods of detecting and / or diagnosing neurological disorders, methods of detecting and / or diagnosing pathological aggregation of tau protein (e.g., in the central nervous system), methods of treating and / or preventing neurological disorders, and methods of promoting the degradation of tau protein by E3 ubiquitin ligase activity in a subject by administering a compound or composition described herein.