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Proceedings of the National Academy of... Jan 2021Recent technological advances have expanded the annotated protein coding content of mammalian genomes, as hundreds of previously unidentified, short open reading frame...
Recent technological advances have expanded the annotated protein coding content of mammalian genomes, as hundreds of previously unidentified, short open reading frame (ORF)-encoded peptides (SEPs) have now been found to be translated. Although several studies have identified important physiological roles for this emerging protein class, a general method to define their interactomes is lacking. Here, we demonstrate that genetic incorporation of the photo-crosslinking noncanonical amino acid AbK into SEP transgenes allows for the facile identification of SEP cellular interaction partners using affinity-based methods. From a survey of seven SEPs, we report the discovery of short ORF-encoded histone binding protein (SEHBP), a conserved microprotein that interacts with chromatin-associated proteins, localizes to discrete genomic loci, and induces a robust transcriptional program when overexpressed in human cells. This work affords a straightforward method to help define the physiological roles of SEPs and demonstrates its utility by identifying SEHBP as a short ORF-encoded transcription factor.
Topics: Amino Acid Sequence; Animals; Cattle; Chromatin; Diazomethane; Gene Expression Regulation; Genetic Loci; HEK293 Cells; HeLa Cells; Histones; Humans; K562 Cells; Lysine; Mice; Open Reading Frames; Pan troglodytes; Peptides; Protein Binding; Protein Interaction Mapping; Rats; Sequence Alignment; Sequence Homology, Amino Acid; Transcription, Genetic; Transgenes; Ultraviolet Rays
PubMed: 33468658
DOI: 10.1073/pnas.2021943118 -
Molecules (Basel, Switzerland) Feb 2023In materials (polymer) science and medicinal chemistry, heteroaromatic derivatives play the role of the central skeleton in development of novel devices and discovery of... (Review)
Review
In materials (polymer) science and medicinal chemistry, heteroaromatic derivatives play the role of the central skeleton in development of novel devices and discovery of new drugs. On the other hand, (3-trifluoromethyl)phenyldiazirine (TPD) is a crucial chemical method for understanding biological processes such as ligand-receptor, nucleic acid-protein, lipid-protein, and protein-protein interactions. In particular, use of TPD has increased in recent materials science to create novel electric and polymer devices with comparative ease and reduced costs. Therefore, a combination of heteroaromatics and (3-trifluoromethyl)diazirine is a promising option for creating better materials and elucidating the unknown mechanisms of action of bioactive heteroaromatic compounds. In this review, a comprehensive synthesis of (3-trifluoromethyl)diazirine-substituted heteroaromatics is described.
Topics: Photoaffinity Labels; Diazomethane; Chemistry, Pharmaceutical; Proteins; Nucleic Acids
PubMed: 36771073
DOI: 10.3390/molecules28031408 -
Journal of Flow Chemistry 2024Azo compounds find use in many areas of science, displaying crucial properties for important applications as photoconductive organic pigments, fluorescent quenchers,... (Review)
Review
Azo compounds find use in many areas of science, displaying crucial properties for important applications as photoconductive organic pigments, fluorescent quenchers, paints, cosmetics, inks, and in the large and valuable dye industry. Due to the unstable intermediates, and the exothermic and fast reactions used in their synthesis, high value azo compounds are excellent candidates for continuous flow manufacturing. This comprehensive review covers the progress made to date on developing continuous flow systems for azo synthesis and reflects on the main challenges still to be addressed, including scale up, conversion, product purity, and environmental impact. The further development of integrated continuous flow processes has the potential to help tackle these challenges and deliver improved methods for azo compound generation.
PubMed: 38882391
DOI: 10.1007/s41981-024-00307-2 -
Antibiotics (Basel, Switzerland) Dec 2022The drug-resistance problem is widely spread and becoming more common in community-acquired and nosocomial strains of bacteria. Therefore, finding new antimicrobial...
The drug-resistance problem is widely spread and becoming more common in community-acquired and nosocomial strains of bacteria. Therefore, finding new antimicrobial agents remains an important drug target. From this perspective, new derivatives of benzothiazole were synthesized and evaluated for their antimicrobial activity and ability to inhibit the DHPS enzyme. The synthesis was carried out by the reaction of benzothiazole -arylsulphonylhydrazone with -aryl-2-cyano-3-(dimethylamino)acrylamide, -aryl-3-(dimethylamino)prop-2-en-1-one, arylaldehydes or diazonium salt of arylamine derivatives, which led to the formation of -arylsulfonylpyridones (yield 60-70%) and - (yield 50-60%)-(2-(benzo[]thiazole-2-yl)-3-arylacryloyl-4-methylsulfonohydrazide - (yield 60-65%) 4-(benzo[]thiazole-2-yl)-5-aryl-1-pyrazol-3(2)-one - (yield 65-75%), and -(2-(benzo[]thiazol-2-yl)-2-(2-arylhydrazono)acetyl)-4-arylsulfonohydrazide - (yield 85-70%). The antimicrobial evaluations resulted into a variety of microbial activities against the tested strains. Most compounds showed antimicrobial activity against with an MIC range of 0.025 to 2.609 mM. The most active compound, , exhibited superior activity against the strain with an of MIC 0.025 mM among all tested compounds, outperforming both standard drugs ampicillin and sulfadiazine. The physicochemical-pharmacokinetic properties of the synthesized compounds were studied, and it was discovered that some compounds do not violate rule of five and have good bioavailability and drug-likeness scores. The five antimicrobial potent compounds with good physicochemical-pharmacokinetic properties were then examined for their inhibition of DHPS enzyme. According to the finding, three compounds, -, had IC values comparable to the standard drug and revealed that compound was the most active compound with an IC value of 7.85 μg/mL, which is comparable to that of sulfadiazine (standard drug) with an IC value of 7.13 μg/mL. A docking study was performed to better understand the interaction of potent compounds with the binding sites of the DHPS enzyme, which revealed that compounds - are linked by two arene-H interactions with Lys220 within the PABA pocket.
PubMed: 36551457
DOI: 10.3390/antibiotics11121799 -
Current Protocols Jul 2021This protocol enables identification of the interaction partners of O-GlcNAcylated proteins. The method involves the introduction of the diazirine photocrosslinker onto...
This protocol enables identification of the interaction partners of O-GlcNAcylated proteins. The method involves the introduction of the diazirine photocrosslinker onto the O-GlcNAc modification within living cells. The photocrosslinker is activated by UV light to yield covalent crosslinking between O-GlcNAcylated proteins and neighboring molecules. The binding partners can be further characterized by immunoblot or proteomics mass spectrometry methods. The benefits of using the photocrosslinker include the capacity to trap low-affinity binding interactions and the ability to selectively target the interaction partners of the O-GlcNAcylated form of the protein of interest. © 2021 Wiley Periodicals LLC. Basic Protocol 1: In-cell production and crosslinking of O-GlcNDAzylated proteins Basic Protocol 2: Immunoblot analysis to assess O-GlcNDAz crosslinking Support Protocol: Detection of UDP-GlcNDAz from cell lysates.
Topics: Acetylglucosamine; Diazomethane; Mass Spectrometry; Protein Processing, Post-Translational; Proteins
PubMed: 34288588
DOI: 10.1002/cpz1.201 -
Molecules (Basel, Switzerland) Jul 2016The synthesis of diazonium salts is historically an important transformation extensively utilized in dye manufacture. However the highly reactive nature of the diazonium...
The synthesis of diazonium salts is historically an important transformation extensively utilized in dye manufacture. However the highly reactive nature of the diazonium functionality has additionally led to the development of many new reactions including several carbon-carbon bond forming processes. It is therefore highly desirable to determine optimum conditions for the formation of diazonium compounds utilizing the latest processing tools such as flow chemistry to take advantage of the increased safety and continuous manufacturing capabilities. Herein we report a series of flow-based procedures to prepare diazonium salts for subsequent in-situ consumption.
Topics: Chemistry Techniques, Synthetic; Diazonium Compounds; Solid-Phase Synthesis Techniques; Temperature
PubMed: 27428944
DOI: 10.3390/molecules21070918 -
Biomacromolecules May 2018Bioadhesives are a current unmet clinical need for mending of blood contacting soft tissues without inducing thrombosis. Recent development of carbene precursor...
Bioadhesives are a current unmet clinical need for mending of blood contacting soft tissues without inducing thrombosis. Recent development of carbene precursor bioadhesives with the advantages of on-demand curing, tuneable modulus, and wet adhesion have been synthesized by grafting diazirine onto poly (amidoamine) (PAMAM-G5) dendrimers. Herein, the structure activity relationships of platelet adhesion and activation is evaluated for the first time on the cured PAMAM-g-diazirine bioadhesives. Three strategies were employed to prevent healthy human donor platelets from adhering and activating on light-cured bioadhesive surfaces: (1) Attenuation of cationic surface charge, (2) antifouling composites by incorporating heparin and alginate in uncured formulation, and (3) heparin wash of cured bioadhesive surface. Topographical imaging of cured and ethanol dehydrated bioadhesive surfaces was used to quantify the adhered and activated platelets with scanning electron microscopy, whose resolution allowed identification of round senescent, short dendritic, and long dendritic platelets. Cured surfaces of PAMAM-g-diazirine (15%) had 10300 ± 500 adhered platelets mm with 99.7% activation into short/long dendritic cells. Reduction of primary amines by higher degree of diazirine grafting or capping of free amines by acetylation reduces platelet adherence (2400 ± 200 vs 3000 ± 300, respectively). Physical incorporation of heparin and alginate in the formulations reduced the activated platelet; 1300 ± 300 and 300 ± 50, activated platelets mm, in comparison with additive free adhesive formulation. Similarly, heparin rinse of the surface of additive free bioadhesive reduced the activated platelet to platelets of heparin composites at 600 ± 100 platelets mm. PAMAM-g-diazirine (15%) bioadhesive retained the photocured mechanical properties and lap shear adhesion despite the addition of heparin and alginate additives.
Topics: Adhesives; Blood Platelets; Cell Adhesion; Cross-Linking Reagents; Diazomethane; Fibrinolytic Agents; Humans; Hydrogels; Methane; Polyamines
PubMed: 29425441
DOI: 10.1021/acs.biomac.8b00074 -
Molecules (Basel, Switzerland) May 2020The use of light-activated chemical probes to study biological interactions was first discovered in the 1960s, and has since found many applications in studying diseases... (Review)
Review
The use of light-activated chemical probes to study biological interactions was first discovered in the 1960s, and has since found many applications in studying diseases and gaining deeper insight into various cellular mechanisms involving protein-protein, protein-nucleic acid, protein-ligand (drug, probe), and protein-co-factor interactions, among others. This technique, often referred to as photoaffinity labelling, uses radical precursors that react almost instantaneously to yield spatial and temporal information about the nature of the interaction and the interacting partner(s). This review focuses on the recent advances in chemical biology in the use of benzophenones and diazirines, two of the most commonly known light-activatable radical precursors, with a focus on the last three years, and is intended to provide a solid understanding of their chemical and biological principles and their applications.
Topics: Benzophenones; Diazomethane; Photoaffinity Labels; Photochemistry
PubMed: 32414020
DOI: 10.3390/molecules25102285 -
Molecules (Basel, Switzerland) Jan 2022A family of oxazaborines, diazaborinones, triazaborines, and triazaborinones was prepared by reaction of polarized ethylenes, such as β-enaminoamides, with...
A family of oxazaborines, diazaborinones, triazaborines, and triazaborinones was prepared by reaction of polarized ethylenes, such as β-enaminoamides, with 4-methylbenzenediazonium tetraphenylborates. The reaction conditions (stirring in CHCl at room temperature (Method A) or stirring with CHCOONa in CHCl at room temperature (Method B) or refluxing in the CHCl/toluene mixture (Method C)) controlled the formation and relative content of these compounds in the reaction mixtures from one to three products. Substituted oxazaborines gradually rearranged into diazaborinones at 250 °C. The prepared compounds were characterized by H NMR, C NMR, IR, and UV-Vis spectroscopy, HRMS, or microanalysis. The structure of individual compounds was confirmed by B NMR, N NMR, 1D NOESY, and X-ray analysis. The mechanism of reaction of enaminoamides with 4-methylbenzenediazonium tetraphenylborate was proposed.
PubMed: 35056681
DOI: 10.3390/molecules27020367 -
ACS Chemical Biology Feb 2021Protein poly-ADP-ribosylation (PARylation) is a heterogeneous and dynamic post-translational modification regulated by various writers, readers, and erasers. It...
Protein poly-ADP-ribosylation (PARylation) is a heterogeneous and dynamic post-translational modification regulated by various writers, readers, and erasers. It participates in a variety of biological events and is involved in many human diseases. Currently, tools and technologies have yet to be developed for unambiguously defining readers and erasers of individual PARylated proteins or cognate PARylated proteins for known readers and erasers. Here, we report the generation of a bifunctional nicotinamide adenine dinucleotide (NAD) characterized by diazirine-modified adenine and clickable ribose. By serving as an excellent substrate for poly-ADP-ribose polymerase 1 (PARP1)-catalyzed PARylation, the generated bifunctional NAD enables photo-cross-linking and enrichment of PARylation-dependent interacting proteins for proteomic identification. This bifunctional NAD provides an important tool for mapping cellular interaction networks centered on protein PARylation, which are essential for elucidating the roles of PARylation-based signals or activities in physiological and pathophysiological processes.
Topics: Azides; Click Chemistry; Cross-Linking Reagents; Diazomethane; HEK293 Cells; Humans; NAD; Poly (ADP-Ribose) Polymerase-1; Poly ADP Ribosylation; Protein Processing, Post-Translational; Proteome; Proteomics; Ultraviolet Rays
PubMed: 33524253
DOI: 10.1021/acschembio.0c00937