2.3 CuAAC in Protein Conjugation

2022 ◽  
Author(s):  
A. La Venia ◽  
A. Kovalová ◽  
M. Vrabel
Keyword(s):  
Protein Conjugation ◽  
Reaction Conditions ◽  
Protein Conjugates ◽  
Cuaac Reaction

AbstractThis chapter summarizes the use of the copper-catalyzed azide–alkyne cycloaddition (CuAAC) reaction in the synthesis of peptide and protein conjugates. The different reaction conditions used for construction of the conjugates and their application in various disciplines are covered. Synthetic strategies for the introduction of the click groups (azide or alkyne) into the peptide backbones are included as well.

scholarly journals Characterization of Hapten-Protein Conjugates: Antibody Generation and Immunoassay Development for Chlorophenoxyacetic Acid Pesticides

2009 ◽  
Vol 92 (6) ◽  
pp. 1773-1779 ◽  
Author(s):  
Robin C Boro ◽  
K Vikas Singh ◽  
C Raman Suri
Keyword(s):  
Inhibitory Concentration ◽  
Dichlorophenoxyacetic Acid ◽  
Protein Conjugation ◽  
Protein Conjugates ◽  
Highly Sensitive ◽  
Ic50 Values ◽  
Chlorophenoxyacetic Acid ◽  
2,4 Dichlorophenoxyacetic Acid

Abstract The generation of specific and sensitive antibodies against small molecules is greatly dependent upon the characteristics of the hapten-protein conjugates. In this study, we report a new fluorescence-based method for the characterization of hapten-protein conjugates. The method is based on an effect promoted by hapten-protein conjugation density upon the fluorescence intensity of the intrinsic tryptophan chromophore molecules of the protein. The proposed methodology is applied to quantify the hapten-protein conjugation density for two different chlorophenoxyacetic acid pesticides, 2,4-dichlorophenoxyacetic acid (2,4-D) and 2,4-dichlorophenoxybutyric acid (2,4-DB), coupled to carrier protein. Highly sensitive anti-2,4-D and anti-2,4-DB antibodies were obtained using these well-characterized hapten-protein conjugates. The generated antibodies were used in an immunoassay format demonstrating inhibitory concentration (IC50) values equal to 30 and 7 ng/mL for 2,4-D and 2,4-DB, respectively. Linearity was observed in the concentration range between 0.1500 ng/mL with LODs around 4 and 3 ng/mL for 2,4-D and 2,4-DB, respectively, in standard water samples. The proposed method was successfully applied for the determination of the extent of hapten-protein conjugation to produce specific antibodies for immunoassay development against pesticides.

scholarly journals Competitive formation of homocircuit [3]rotaxanes in synthetically useful yields in the bipyridine-mediated active template CuAAC reaction

2015 ◽  
Vol 6 (4) ◽  
pp. 2398-2404 ◽  
Author(s):  
Edward A. Neal ◽  
Stephen M. Goldup
Keyword(s):  
Quantitative Yield ◽  
Reaction Conditions ◽  
Cuaac Reaction

We demonstrate that, depending on reaction conditions, [2]rotaxanes are produced in essentially quantitative yield in the AT-CuAAC reaction regardless of macrocycle size, and hard to access doubly threaded [3]rotaxanes can be synthesised in up to 50% isolated yield in a four component coupling step.

scholarly journals An eco-compatible strategy for the diversity-oriented synthesis of macrocycles exploiting carbohydrate-derived building blocks

2017 ◽  
Vol 13 ◽  
pp. 1106-1118 ◽  
Author(s):  
Sushil K Maurya ◽  
Rohit Rana
Keyword(s):  
Building Blocks ◽  
Ring Size ◽  
Macrocyclic Compounds ◽  
Reaction Conditions ◽  
Structural Class ◽  
Diversity Oriented Synthesis ◽  
Cuaac Reaction ◽  
Standard Reaction

An efficient, eco-compatible diversity-oriented synthesis (DOS) approach for the generation of library of sugar embedded macrocyclic compounds with various ring size containing 1,2,3-triazole has been developed. This concise strategy involves the iterative use of readily available sugar-derived alkyne/azide–alkene building blocks coupled through copper catalyzed azide–alkyne cycloaddition (CuAAC) reaction followed by pairing of the linear cyclo-adduct using greener reaction conditions. The eco-compatibility, mild reaction conditions, greener solvents, easy purification and avoidance of hazards and toxic solvents are advantages of this protocol to access this important structural class. The diversity of the macrocycles synthesized (in total we have synthesized 13 macrocycles) using a set of standard reaction protocols demonstrate the potential of the new eco-compatible approach for the macrocyclic library generation.

scholarly journals Exploiting Protein N-Terminus for Site-Specific Bioconjugation

Molecules ◽  
2021 ◽  
Vol 26 (12) ◽  
pp. 3521
Author(s):  
Lucia De Rosa ◽  
Rossella Di Stasi ◽  
Alessandra Romanelli ◽  
Luca Domenico D’Andrea
Keyword(s):  
Protein Molecule ◽  
Amine Group ◽  
Reactive Site ◽  
Reaction Conditions ◽  
Selective Labeling ◽  
Site Specific ◽  
Protein Conjugates ◽  
Protein Molecules ◽  
N Terminus

Although a plethora of chemistries have been developed to selectively decorate protein molecules, novel strategies continue to be reported with the final aim of improving selectivity and mildness of the reaction conditions, preserve protein integrity, and fulfill all the increasing requirements of the modern applications of protein conjugates. The targeting of the protein N-terminal alpha-amine group appears a convenient solution to the issue, emerging as a useful and unique reactive site universally present in each protein molecule. Herein, we provide an updated overview of the methodologies developed until today to afford the selective modification of proteins through the targeting of the N-terminal alpha-amine. Chemical and enzymatic strategies enabling the selective labeling of the protein N-terminal alpha-amine group are described.

Carbohydrate Protein Conjugates (CPC): The Design of New Materials to Stabilize Enzymes

1990 ◽  
Vol 218 ◽  
Author(s):  
Tara G. Hill ◽  
Peng Wang ◽  
Lynn M. Oehlert ◽  
Michael E. Huston ◽  
Charles A. Wartchow ◽  
...  
Keyword(s):  
Organic Synthesis ◽  
Covalent Attachment ◽  
Ion Exchange Resins ◽  
Chemical Derivatization ◽  
New Materials ◽  
Reaction Conditions ◽  
Protein Conjugates ◽  
Exchange Resins ◽  
Polymeric Supports ◽  
Protein Covalent

Intense efforts have been directed at the stabilization of proteins because of their potential uses in organic synthesis, diagnostics, and the pharmaceutical industry. These efforts have resulted in a number of methods to stabilize enzymes including adsorbtion on inert supports or ion exchange resins, entrapment within a gel (with or without crosslinking of the gel or protein), covalent attachment to beads or polymeric supports, inclusion in micelles, chemical derivatization of the protein and mutagenesis. However, these methods do not provide a general approach to solving the problem of protein stability. We believed that the multi-site attachment of a carbohydrate-based macromolecule to the surface of a protein would provide structural stability and a water-like microenvironment for the protein under harsh reaction conditions.

scholarly journals A Hybrid Detection Method Based on Peroxidase-mediated Signal Amplification and Click Chemistry for Highly Sensitive Background-free Immunofluorescent Staining

2019 ◽  
Vol 67 (10) ◽  
pp. 771-782
Author(s):  
Stanislav A. Antonov ◽  
Ekaterina V. Novosadova ◽  
Andrey G. Kobylansky ◽  
Vyacheslav Z. Tarantul ◽  
Igor A. Grivennikov
Keyword(s):  
Signal Amplification ◽  
Detection Method ◽  
Detection Threshold ◽  
Immunofluorescent Staining ◽  
Reaction Conditions ◽  
Tissue Sections ◽  
Cuaac Reaction ◽  
Highly Sensitive ◽  
Hybrid Detection ◽  
Catalyzed Reporter Deposition

The copper-catalyzed azide-alkyne cycloaddition (CuAAC) reaction is increasingly used for detection of various macromolecules and metabolites in biological samples. Here, we present a detailed analysis of the CuAAC reaction conditions in cells and tissue sections. Using the optimized CuAAC conditions, we have devised a highly sensitive immunostaining technique, based on the tyramide signal amplification/catalyzed reporter deposition (TSA/CARD) method with a novel alkyne tyramide substrate. The described method offers improved detection threshold compared to conventional immunofluorescent staining and produces significantly lower non-specific background than TSA/CARD with fluorescent tyramides.

Copper-catalyzed alkyne-azide cycloaddition for the functionalization of fullerene building blocks

2011 ◽  
Vol 84 (4) ◽  
pp. 1027-1037 ◽  
Author(s):  
Jean-François Nierengarten
Keyword(s):  
Chemical Transformation ◽  
Building Blocks ◽  
Reaction Conditions ◽  
Cuaac Reaction

In this paper, we report our ongoing progress in the preparation of fullerene-azide or fullerene-alkyne building blocks, allowing their further chemical transformation under the copper-catalyzed alkyne-azide cycloaddition (CuAAC) reaction conditions.

Undecagold labeling of tRNA

1991 ◽  
Vol 49 ◽  
pp. 44-45
Author(s):  
James F. Hainfeld ◽  
Kyra M. Alford ◽  
Mathias Sprinzl ◽  
Valsan Mandiyan ◽  
Santa J. Tumminia ◽  
...  
Keyword(s):  
High Resolution ◽  
Transmission Electron Micrograph ◽  
Anticodon Loop ◽  
Electron Micrograph ◽  
Reaction Conditions ◽  
Scanning Transmission Electron ◽  
Transmission Electron ◽  
Scanning Transmission

The undecagold (Au11) cluster was used to covalently label tRNA molecules at two specific ribonucleotides, one at position 75, and one at position 32 near the anticodon loop. Two different Au11 derivatives were used, one with a monomaleimide and one with a monoiodacetamide to effect efficient reactions.The first tRNA labeled was yeast tRNAphe which had a 2-thiocytidine (s2C) enzymatically introduced at position 75. This was found to react with the iodoacetamide-Aun derivative (Fig. 1) but not the maleimide-Aun (Fig. 2). Reaction conditions were 37° for 16 hours. Addition of dimethylformamide (DMF) up to 70% made no improvement in the labeling yield. A high resolution scanning transmission electron micrograph (STEM) taken using the darkfield elastically scattered electrons is shown in Fig. 3.

Cardiological Biopharmaceuticals in the Conception of Drug Targeting Delivery: Practical Results and Research Perspectives

Acta Naturae ◽  
2012 ◽  
Vol 4 (3) ◽  
pp. 72-81 ◽  
Author(s):  
A. V. Maksimenko
Keyword(s):  
Cell Adhesion Molecules ◽  
Drug Targeting ◽  
Interventional Procedures ◽  
Target Localization ◽  
Clinical Use ◽  
Protein Conjugates ◽  
Research Perspectives ◽  
Significant Interest ◽  
Targeting Delivery ◽  
Molecular Sizes

The results of the clinical use of thrombolytic and antithrombotic preparations developed on the basis of protein conjugates obtained within the framework of the conception of drug targeting delivery in the organism are considered. A decrease has been noted in the number of biomedical projects focused on these derivatives as a result of various factors: the significant depletion of financial and organizational funds, the saturation of the pharmaceutical market with preparations of this kind, and the appearance of original means for interventional procedures. Factors that actively facilitate the conspicuous potentiation of the efficacy of bioconjugates were revealed: the biomedical testing of protein domains and their selected combinations, the optimization of molecular sizes for the bioconjugates obtained, the density of target localization, the application of cell adhesion molecules as targets, and the application of connected enzyme activities. Enzyme antioxidants and the opportunity for further elaboration of the drug delivery conception via the elucidation and formation of therapeutic targets for effective drug reactions by means of pharmacological pre- and postconditioning of myocardium arouse significant interest.

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