Multifunctional streptavidin–biotin conjugates with precise stoichiometries

Xu, Dongdong; Wegner, Seraphine V.

doi:10.1039/d0sc01589j

Cited by 13 publications

(13 citation statements)

References 23 publications

(29 reference statements)

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“…Tetravalent biotin–streptavidin ( STV ) binding provides a modular high-affinity strategy for protein bioconjugation. − A wide range of molecular and macromolecular systems can be biotinylated, making this a highly versatile platform for bioconjugation. , We report here the use of biotin–streptavidin assemblies to provide cytosolic delivery of proteins into cells. In this system, biotinylated oligo(glutamate) ( b-E20 ) and biotinylated proteins were first bound to STV (Figure ).…”

Section: Introductionmentioning

confidence: 99%

Cytosolic Protein Delivery Using Modular Biotin–Streptavidin Assembly of Nanocomposites

et al. 2022

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Current strategies for the delivery of proteins into cells face general challenges of endosomal entrapment and concomitant degradation of protein cargo. Efficient delivery directly to the cytosol overcomes this obstacle: we report here the use of biotin–streptavidin tethering to provide a modular approach to the generation of nanovectors capable of a cytosolic delivery of biotinylated proteins. This strategy uses streptavidin to organize biotinylated protein and biotinylated oligo(glutamate) peptide into modular complexes that are then electrostatically self-assembled with a cationic guanidinium-functionalized polymer. The resulting polymer–protein nanocomposites demonstrate efficient cytosolic delivery of six biotinylated protein cargos of varying size, charge, and quaternary structure. Retention of protein function was established through efficient cell killing via delivery of the chemotherapeutic enzyme granzyme A. This platform represents a versatile and modular approach to intracellular delivery through the noncovalent tethering of multiple components into a single delivery vector.

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Section: Introductionmentioning

confidence: 99%

Cytosolic Protein Delivery Using Modular Biotin–Streptavidin Assembly of Nanocomposites

et al. 2022

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“…This is in contrast to the successful immobilization of streptavidin as demonstrated earlier, but not surprising considering the relatively poor binding of the mannose-ConA couple as compared to the biotin−streptavidin couple (K d = 2.89 × 10 −6 M and 1 × 10 −14 M, respectively). 50,51 Although from the fluorescence microscopy analysis, it was evident that, for the surface obtained using mannose modification of the G1-alkene, a slightly higher amount of ConA binding took place and it did not appear to be effective. To understand if the crowding of functional groups leads to this lack of enhanced protein binding, we decided to dilute the mannose clusters by adding AIBN during the attachment of the alkene units.…”

Section: ■ Results and Discussionmentioning

confidence: 99%

“Clickable” Polymer Brush Interfaces: Tailoring Monovalent to Multivalent Ligand Display for Protein Immobilization and Sensing

et al. 2022

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Facile and effective functionalization of the interface of polymer-coated surfaces allows one to dictate the interaction of the underlying material with the chemical and biological analytes in its environment. Herein, we outline a modular approach that would enable installing a variety of “clickable” handles onto the surface of polymer brushes, enabling facile conjugation of various ligands to obtain functional interfaces. To this end, hydrophilic anti-biofouling poly(ethylene glycol)-based polymer brushes are fabricated on glass-like silicon oxide surfaces using reversible addition–fragmentation chain transfer (RAFT) polymerization. The dithioester group at the chain-end of the polymer brushes enabled the installation of azide, maleimide, and terminal alkene functional groups, using a post-polymerization radical exchange reaction with appropriately functionalized azo-containing molecules. Thus, modified polymer brushes underwent facile conjugation of alkyne or thiol-containing dyes and ligands using alkyne–azide cycloaddition, Michael addition, and radical thiol–ene conjugation, respectively. Moreover, we demonstrate that the radical exchange approach also enables the installation of multivalent motifs using dendritic azo-containing molecules. Terminal alkene groups containing dendrons amenable to functionalization with thiol-containing molecules using the radical thiol–ene reaction were installed at the interface and subsequently functionalized with mannose ligands to enable sensing of the Concanavalin A lectin.

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“…Next, the synthesized Fe-N x SANs were treated with N-(3-dimethylamino propyl)-N′-ethylcarbodiimide hydrochloride (EDC) and N-hydroxysuccinimide (NHS), then modified with SA to bind biotinylated A β 1-40 antibody, which is proved by the Fourier transform infrared spectra successfully (Figure S1 ) [ 32 ]. Thereinto, the biotin can react with SA-conjugated labels, forming the strongest known noncovalent interaction between a protein and a ligand [ 33 ]. Notably, the interaction is rapid and maintains being robust in extreme conditions of pH and temperature.…”

Section: Resultsmentioning

confidence: 99%

Single-Atom Nanozymes Linked Immunosorbent Assay for Sensitive Detection of A β 1-40: A Biomarker of Alzheimer’s Disease

et al. 2020

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Single-atom nanozymes (SANs) possess unique features of maximum atomic utilization and present highly assembled enzyme-like structure and remarkable enzyme-like activity. By introducing SANs into immunoassay, limitations of ELISA such as low stability of horseradish peroxidase (HRP) can be well addressed, thereby improving the performance of the immunoassays. In this work, we have developed novel Fe-N-C single-atom nanozymes (Fe-Nx SANs) derived from Fe-doped polypyrrole (PPy) nanotube and substituted the enzymes in ELISA kit for enhancing the detection sensitivity of amyloid beta 1-40. Results indicate that the Fe-Nx SANs contain high density of single-atom active sites and comparable enzyme-like properties as HRP, owing to the maximized utilization of Fe atoms and their abundant active sites, which could mimic natural metalloproteases structures. Further designed SAN-linked immunosorbent assay (SAN-LISA) demonstrates the ultralow limit of detection (LOD) of 0.88 pg/mL, much more sensitive than that of commercial ELISA (9.98 pg/mL). The results confirm that the Fe-Nx SANs can serve as a satisfactory replacement of enzyme labels, which show great potential as an ultrasensitive colorimetric immunoassay.

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Multifunctional streptavidin–biotin conjugates with precise stoichiometries

Abstract: Multifunctional streptavidin-biotin conjugates with defined stoichiometry and number of open binding pockets provide molecularly precise alternatives to the statistical mixture of products that typically forms.

Cited by 13 publications

References 23 publications

Cytosolic Protein Delivery Using Modular Biotin–Streptavidin Assembly of Nanocomposites

Cytosolic Protein Delivery Using Modular Biotin–Streptavidin Assembly of Nanocomposites

“Clickable” Polymer Brush Interfaces: Tailoring Monovalent to Multivalent Ligand Display for Protein Immobilization and Sensing

Single-Atom Nanozymes Linked Immunosorbent Assay for Sensitive Detection of A β 1-40: A Biomarker of Alzheimer’s Disease

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