A “Catch‐and‐Release” Protocol for Alkyne‐Tagged Molecules Based on a Resin‐Bound Cobalt Complex for Peptide Enrichment in Aqueous Media

The development of new and mild protocols for the specific enrichment of biomolecules is of significant interest from the perspective of chemical biology. A cobalt–phosphine complex immobilised on a solid‐phase resin has been found to selectively bind to a propargyl carbamate tag, that is, “catch”, under dilute aqueous conditions (pH 7) at 4 °C. Upon acidic treatment of the resulting resin‐bound alkyne–cobalt complex, the Nicholas reaction was induced to “release” the alkyne‐tagged molecule from the resin as a free amine. Model studies revealed that selective enrichment of the alkyne‐tagged molecule could be achieved with high efficiency at 4 °C. The proof‐of‐concept was applied to an alkyne‐tagged amino acid and dipeptide. Studies using an alkyne‐tagged dipeptide proved that this protocol is compatible with various amino acids bearing a range of functionalities in the side‐chain. In addition, selective enrichment and detection of an amine derived from the “catch and release” of an alkyne‐tagged dipeptide in the presence of various peptides has been accomplished under highly dilute conditions, as determined by mass spectrometry.     

Potassium Acyltrifluoroborate (KAT) Ligations are Orthogonal to Thiol‐Michael and SPAAC Reactions: Covalent Dual Immobilization of Proteins onto Synthetic PEG Hydrogels

The covalent immobilization of peptides, proteins, and other biomolecules to hydrogels provides a biologically mimicking environment for cell and tissue growth. Bioorthogonal chemical reactions can serve as a tool for this, but the paucity of such reactions and mutual incompatibilities limits the number of distinct molecules that can be introduced. We now report that the potassium acyltrifluoroborate (KAT ) amide‐forming ligation is orthogonal to both thiol‐Michael and strain promoted azide alkyne cycloadditions (SPAAC ) and the requisite functional groups – KAT s and hydroxylamines – are stable and compatible to hydrogel formation, protein modification, and post‐assembly immobilization of biomolecules onto hydrogels. In combination these ligations enables stepwise covalent protein immobilization of multiple BSA ‐derivatives onto the hydrogel scaffold regardless of the order of addition.     

Azidopropylvinylsulfonamide as a New Bifunctional Click Reagent for Bioorthogonal Conjugations: Application for DNA–Protein Cross‐Linking

N‐(3‐Azidopropyl)vinylsulfonamide was developed as a new bifunctional bioconjugation reagent suitable for the cross‐linking of biomolecules through copper(I)‐catalyzed azide–alkyne cycloaddition and thiol Michael addition reactions under biorthogonal conditions. The reagent is easily clicked to an acetylene‐containing DNA or protein and then reacts with cysteine‐containing peptides or proteins to form covalent cross‐links. Several examples of bioconjugations of ethynyl‐ or octadiynyl‐modified DNA with peptides, p53 protein, or alkyne‐modified human carbonic anhydrase with peptides are given.     

Click‐chemistry of polymersomes on nanoporous polymeric surfaces

A facile click chemistry method of immobilizing surface‐functionalized polymer vesicles on casted polymeric PAN substrates is described. Microporous PAN membranes were subjected to hydrochloric acid hydrolysis to obtain surface carboxylates. The carboxylic groups were activated with EDC/NHS‐solution and were then reacted with propargylamine to introduce alkyne groups for CuAAC reactions. The alkyne functionality of the modified membrane surface was verified by reaction with an azide functional click dye both before and after the immobilization of azide‐functionalized ABA vesicles. The efficient postfunctionalization of the membrane with alkyne allowed quantitative coverage of the membrane surface with a polymersome monolayer, as confirmed by immobilization of polymerzomes loaded with a fluorescent dye. Polymersome monolayers immobilized on alkyne functionalized PAN‐membranes were characterized by cryo‐SEM and monolayers were confirmed by atom force microscopy. These methods opens up new avenues for preparing membrane based filtration and sensor technologies. © 2016 Wiley Periodicals, Inc. J. Polym. Sci., Part A: Polym. Chem. 2016 , 54, 2032–2039     

Photo‐ and pH‐Responsive Polycarbonate Block Copolymer Prodrug Nanomicelles for Controlled Release of Doxorubicin

Photo/pH dual‐responsive amphiphilic diblock copolymers with alkyne functionalized pendant o‐nitrobenzyl ester group are synthesized using poly(ethylene glycol) as a macroinitiator. The pendant alkynes are functionalized as aldehyde groups by the azide‐alkyne Huisgen cycloaddition. The anticancer drug doxorubicin (DOX) molecules are then covalently conjugated through acid‐sensitive Schiff‐base linkage. The resultant prodrug copolymers self‐assemble into nanomicelles in aqueous solution. The prodrug nanomicelles have a well‐defined morphology with an average size of 20–40 nm. The dual‐stimuli are applied individually or simultaneously to study the release behavior of DOX. Under UV light irradiation, nanomicelles are disassembled due to the ONB ester photocleavage. The light‐controlled DOX release behavior is demonstrated using fluorescence spectroscopy. Due to the pH‐sensitive imine linkage the DOX molecules are released rapidly from the nanomicelles at the acidic pH of 5.0, whereas only minimal amount of DOX molecules is released at the pH of 7.4. The DOX release rate is tunable by applying the dual‐stimuli simultaneously. In vitro studies against colon cancer cells demonstrate that the nanomicelles show the efficient cellular uptake and the intracellular DOX release, indicating that the newly designed copolymers with dual‐stimuli‐response have significant potential applications as a smart nanomedicine against cancer.     

Live‐Cell Stimulated Raman Scattering Imaging of Alkyne‐Tagged Biomolecules

Alkynes can be metabolically incorporated into biomolecules including nucleic acids, proteins, lipids, and glycans. In addition to the clickable chemical reactivity, alkynes possess a unique Raman scattering within the Raman‐silent region of a cell. Coupling this spectroscopic signature with Raman microscopy yields a new imaging modality beyond fluorescence and label‐free microscopies. The bioorthogonal Raman imaging of various biomolecules tagged with an alkyne by a state‐of‐the‐art Raman imaging technique, stimulated Raman scattering (SRS) microscopy, is reported. This imaging method affords non‐invasiveness, high sensitivity, and molecular specificity and therefore should find broad applications in live‐cell imaging.     

In‐column preparation of a brush‐type chiral stationary phase using click chemistry and a silica monolith

Brush‐type chiral stationary phases (CSP) have been prepared both from a silica monolith and, separately, from 10 μm porous silica beads via a process of in‐column modification including attachment of the chiral selector via copper‐catalyzed azide–alkyne cycloaddition. Azide functionalities were first introduced on the pore surface of each type of support by reaction with 3‐(azidopropyl)trimethoxysilane, followed by immobilization of a proline‐derived chiral selector containing an alkyne moiety. This functionalization reaction was carried out in dimethylformamide (DMF) in the presence of catalytic amounts of copper (I) iodide. The separation performance of these triazole linked stationary phases was demonstrated in enantioseparations of four model analytes, which afforded separation factors as high as 11.4.     

Pyridine‐Enhanced Head‐to‐Tail Dimerization of Terminal Alkynes by a Rhodium–N‐Heterocyclic‐Carbene Catalyst

A general regioselective rhodium‐catalyzed head‐to‐tail dimerization of terminal alkynes is presented. The presence of a pyridine ligand (py) in a Rh–N‐heterocyclic‐carbene (NHC) catalytic system not only dramatically switches the chemoselectivity from alkyne cyclotrimerization to dimerization but also enhances the catalytic activity. Several intermediates have been detected in the catalytic process, including the π‐alkyne‐coordinated Rh^I species [RhCl(NHC)(η²‐HC?CCH₂Ph)(py)] ( 3 ) and [RhCl(NHC){η²‐C(tBu)?C(E)CH?CHtBu}(py)] ( 4 ) and the Rh^III–hydride–alkynyl species [RhClH{? C?CSi(Me)₃}(IPr)(py)₂] ( 5 ). Computational DFT studies reveal an operational mechanism consisting of sequential alkyne C? H oxidative addition, alkyne insertion, and reductive elimination. A 2,1‐hydrometalation of the alkyne is the more favorable pathway in accordance with a head‐to‐tail selectivity.     

Precision PEGylated Polymers Obtained by Sequence‐Controlled Copolymerization and Postpolymerization Modification

Copolymers containing water‐soluble poly(ethylene glycol) (PEG) side chains and precisely controlled functional microstructures were synthesized by sequence‐controlled copolymerization of donor and acceptor comonomers, that is, styrene derivatives and N‐substituted maleimides. Two routes were compared for the preparation of these structures: a) the direct use of a PEG–styrene macromonomer as a donor comonomer, and b) the use of an alkyne‐functionalized styrenic comonomer, which was PEGylated by copper‐catalyzed alkyne–azide cycloaddition after polymerization. The latter method was found to be the most versatile and enabled the synthesis of high‐precision copolymers. For example, PEGylated copolymers containing precisely positioned fluorescent (e.g. pyrene), switchable (e.g. azobenzene), and reactive functionalities (e.g. an activated ester) were prepared.     

Acyl guanidine functional poly(2‐oxazoline)s as reactive intermediates and stimuli‐responsive materials

The many postpolymerization modification opportunities of biocompatible poly(2‐alkyl/aryl‐2‐oxazoline)s (PAOx), such as thiol–ene/thiol–yne, azide–alkyne cycloadditions, amidation, and transesterification, are one of the most appealing features of this polymer class for its popularity in biomedicine. Inspired by recent reports on guanidine‐catalyzed transesterification and amidation reactions of methyl ester substrates, we explored the use of guanidines as a reactant for the modification of methyl ester functional PAOx, to obtain the respective acyl guanidines. The obtained acyl guanidines functional polymers display reactivity toward α‐haloketones, yielding imidazole functional PAOx. The obtained polymer structures are protonated in a broad pH range, and the acyl guanidine moiety is demonstrated to be a cleavable linker under basic conditions. © 2019 Wiley Periodicals, Inc. J. Polym. Sci., Part A: Polym. Chem. 2019 , 57, 2616–2624     

Surface‐Guided Formation of an Organocobalt Complex

Organocobalt complexes represent a versatile tool in organic synthesis as they are important intermediates in Pauson–Khand, Friedel–Crafts, and Nicholas reactions. Herein, a single‐molecule‐level investigation addressing the formation of an organocobalt complex at a solid–vacuum interface is reported. Deposition of 4,4′‐(ethyne‐1,2‐diyl)dibenzonitrile and Co atoms on the Ag(111) surface followed by annealing resulted in genuine complexes in which single Co atoms laterally coordinated to two carbonitrile groups undergo organometallic bonding with the internal alkyne moiety of adjacent molecules. Alternative complexation scenarios involving fragmentation of the precursor were ruled out by complementary X‐ray photoelectron spectroscopy. According to density functional theory analysis, the complexation with the alkyne moiety follows the Dewar–Chatt–Duncanson model for a two‐electron‐donor ligand where an alkyne‐to‐Co donation occurs together with a strong metal‐to‐alkyne back‐donation.     

Reduction‐cleavable hyperbranched polymers with limited intramolecular cyclization via click chemistry

In this contribution, we present new reduction‐cleavable hyperbranched disulfide bonds‐containing poly(ester triazole)s with limited intramolecular cyclization, which can be synthesized by the Cu(I)‐catalyzed azide–alkyne cycloaddition (CuAAC) of A₂ monomer of dipropargyl 3,3′‐dithiobispropionate and B₃ monomer of tris(hydroxymethyl)ethane tri(4‐azidobutanoate). The hyperbranched poly(ester triazole)s possess numerous terminal groups and weight‐average molecular weight up to 20,400 g mol^?1 with a polydispersity index in the range 1.57–2.17. The CuAAC introduces rigid triazole units into the backbones of hyperbranched poly(ester triazole)s and reduces intramolecular cyclization, which is proved by topological analysis and ¹H NMR spectroscopy. The disulfide bonds on backbones endow the reduction‐cleavable feature to the hyperbranched poly(ester triazole)s at the presence of dithiothreitol. It gives a novel and convenient methodology for the synthesis of reduction‐responsive functional polymer with controlled topologies, and the reduction‐cleavable hyperbranched poly(ester triazole)s with limited intramolecular cyclization are expected to possess potential in the application of stimuli‐responsive anticancer drug nanocarriers. © 2015 Wiley Periodicals, Inc. J. Polym. Sci., Part A: Polym. Chem. 2015 , 53, 2374–2380     

Tracking Surface Glycans on Live Cancer Cells with Single‐Molecule Sensitivity

Using a combination of metabolically labeled glycans, a bioorthogonal copper(I)‐catalyzed azide–alkyne cycloaddition, and the controlled bleaching of fluorescent probes conjugated to azide‐ or alkyne‐tagged glycans, a sufficiently low spatial density of dye‐labeled glycans was achieved, enabling dynamic single‐molecule tracking and super‐resolution imaging of N‐linked sialic acids and O‐linked N‐acetyl galactosamine (GalNAc) on the membrane of live cells. Analysis of the trajectories of these dye‐labeled glycans in mammary cancer cells revealed constrained diffusion of both N‐ and O‐linked glycans, which was interpreted as reflecting the mobility of the glycan rather than to be caused by transient immobilization owing to spatial inhomogeneities on the plasma membrane. Stochastic optical reconstruction microscopy (STORM) imaging revealed the structure of dynamic membrane nanotubes.     

Azide/alkyne‐“click”‐reactions on amino resin materials: An LC‐ESI‐TOF analysis

This article describes the reaction of amino resins with functional molecules using the azide/alkyne‐“click”‐reaction, opening a simple chemical modification of amino resins under aqueous conditions. Alkyne‐modified melamine‐formaldehyde resins are prepared via a direct cocondensation approach using propargylic alcohol (21.6–86.3 mmol) as additive. Subsequently, alkyne‐modified mono‐, bi‐, and trinuclear melamine‐species are identified via LC‐ESI‐TOF methods proving the covalent incorporation of alkyne‐moieties in amounts of up to 3.9 mol %. Subsequent modification of the alkyne‐modified resins was accomplished by reaction of functional azides (octyl azide (1), (azidomethyl)benzene (2), 1‐(6‐azidohexyl) thymine (3), and 4‐azido‐N‐(2,2,6,6‐tetramethylpiperidin‐4‐yl)benzamide (4)) with Cu(I)Br and DIPEA as a base. The formation of triazolyl‐modified MF‐resins was proven by LC‐ESI‐TOF methods, indicating the successful covalent modification of the amino resin with the azides 1 – 4 . © 2010 Wiley Periodicals, Inc. J Polym Sci Part A: Polym Chem, 2010     

V‐shaped graft copolymers via triple click reactions: Diels–alder,copper‐catalyzed azide–alkyne cycloaddition,and nitroxide radical coupling

We report here a simple and universal synthetic pathway covering triple click reactions, Diels–Alder, copper‐catalyzed azide–alkyne cycloaddition (CuAAC), and nitroxide radical coupling (NRC), to prepare well‐defined graft copolymers with V‐shaped side chains. The Diels–Alder click reaction between the furan protected‐maleimide‐terminated poly(ethylene glycol) (PEG) and a trifunctional core ( 1 ) carrying an anthracene, alkyne, and bromide was carried out to yield the corresponding α‐alkyne‐ and α‐bromide‐terminated PEG (PEG‐alkyne/Br) in toluene at 110 °C. Subsequently, the polystyrene or polyoxanorbornene with pendant azide functionality as a main backbone is reacted with the PEG‐alkyne/Br and 2,2,6,6‐tetramethyl‐1‐piperidinyloxy (TEMPO)‐terminated poly(ε‐caprolactone) using the CuAAC and NRC reactions in a one‐pot fashion in N,N′‐dimethylformamide at room temperature to result in the target V‐shaped graft copolymers. © 2013 Wiley Periodicals, Inc. J. Polym. Sci., Part A: Polym. Chem. 2013 , 51, 4667–4674     

Determining the Origin of Rate‐Independent Chemoselectivity in CuAAC Reactions: An Alkyne‐Specific Shift in Rate‐Determining Step

We report a kinetic and spectroscopic analysis of alkyne‐dependent chemoselectivity in the copper‐catalyzed azide–alkyne click (CuAAC) reaction. Studies of six alkyne subtypes reveal that the rate‐determining step (RDS) of an aromatic ynamine class is shifted from acetylide formation to the azide ligation/migratory insertion event allowing chemoselectivity independent of overall rate.     

Development and Characterization of Nickel‐NTA‐Polyaniline Modified Electrodes

The engineered addition of hexa‐histidine sequences to biomolecules such as antibody fragments has been found to be an excellent means of purifying these materials. This tagging methodology has also been extended to its use as a tool for immobilization and orientation of antibodies on transducer surfaces. Polyvinyl sulfonate‐doped polyanilne (PANI/PVS) can be used as a mediator in amperometric biosensors. This short communication looks at the effect of nickel chelate materials and nickel chelation on this conducting polymer and evaluates it as a potential surface for the immobilization of his‐tagged biomolecules. N‐nitrilotriacetic acid (NTA) was doped into the electropolymerized PANI/PVS at a screen‐printed carbon paste electrode. The resulting NTA‐PANI/PVS film was shown to have comparable electrochemical properties of polymer without the chelating agent. When Ni²⁺ was applied to the electrode, the incorporated NTA was found to efficiently chelate the metal ions at the electrode surface.     

A Novel Pd‐Catalysed Annulation Reaction for the Syntheses of Pyrroloindoles and Pyrroloquinolines

Pd‐catalysed annulation reactions between indole derivatives and internal alkyne esters leading to various pyrrolo[1,2‐a]indoles and pyrroloquinolines have been developed. The strategy involves an intermolecular addition of the indole nitrogen on to the internal alkyne ester followed by an intramolecular insertion of a vinyl–palladium complex into the carbonyl group. This method offers a facile and practical approach to pyrrolo[1,2‐a]indoles and pyrroloquinolines.     

One‐Pot synthesis of functional poly(methacrylate) by ATRP and 1,8‐Diazacyclo‐[5,4,0]undec‐7‐ene catalyzed transesterification

A facile one‐pot 1,8‐diazacyclo‐[5,4,0]undec‐7‐ene (DBU) catalyzed transesterification/atom transfer radical polymerization (ATRP) strategy has been successfully developed through the combination of copper/DBU‐catalyzed ATRP and DBU‐catalyzed transesterification reactions. Well‐defined poly(methacrylate)s with various side ester groups have been synthesized by ATRP and transesterification of acyl donor 2,2,2‐trifluoethyl methacrylate with various alcohols, such as benzyl alcohol, n‐butanol, iso‐propyl alcohol, methanol, triethylene glycol monomethyl ether, propargyl alcohol, and 6‐azido‐1‐hexanol by the one‐pot process. Kinetic studies indicate that the ATRP process proceeded in a controllable manner without the interference of the transesterification reactions. Expansion of the binary system to a higher level ternary system has been successfully achieved by the combination of copper(I)‐catalyzed azide–alkyne cycloaddition, transesterification, and ATRP reactions. © 2014 Wiley Periodicals, Inc. J. Polym. Sci., Part A: Polym. Chem. 2014 , 52, 2998–3003     

An Alkyne‐Appended,Click‐Ready PtII Complex with an Unusual Arrangement in the Solid State

To better understand the range of cellular interactions of Pt^II‐based chemotherapeutics, robust and efficient methods to track and analyze Pt targets are needed. A powerful approach is to functionalize Pt^II compounds with alkyne or azide moieties for post‐treatment conjugation through the azide–alkyne cycloaddition (click) reaction. Herein, we report an alkyne‐appended cis‐diamine Pt^II compound, cis‐[Pt(2‐(5‐hexynyl)amido‐1,3‐propanediamine)Cl₂] ( 1 ), the X‐ray crystal structure of which exhibits a combination of unusual radially distributed CH/π(CC) interactions, Pt Pt bonding, and NH:O/NH:Cl hydrogen bonds. In solution, 1 exhibits no Pt alkyne interactions and binds readily to DNA. Subsequent click reactivity with nonfluorescent dansyl azide results in a 70‐fold fluorescence increase. This result demonstrates the potential for this new class of alkyne‐modified Pt compound for the comprehensive detection and isolation of Pt‐bound biomolecules.