Preparation and Characterization of Quantum Dot-Peptide Conjugates Based on Polyhistidine Tags

Krause, Katherine D.; Tsai, Hsin-Yun; Rees, Kelly; Kim, Hyungki; Algar, W. Russ

doi:10.1007/978-1-0716-1617-8_16

Cited by 3 publications

(4 citation statements)

References 97 publications

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“…Proteolysis was tracked ratiometrically via the loss of FRET-sensitized sCy5 emission and the concomitant increase in QD PL emission upon hydrolysis of the substrate and diffusion of the sCy5-labeled terminus away from the QD. The substrate peptide sequences were designed with functional blocks of amino acids (Figure D): (1) an N -terminal hexahistidine sequence, which spontaneously binds to the inorganic surface of QDs; − (2) a polyproline sequence, which was anticipated to form a helical spacer to orient the peptide away from the QD surface; (3) a hydrophilic sequence that incorporated (4) a recognition site for the protease of interest (see the SI for discussion); and (5) a terminal cysteine residue for site-selective labeling with sCy5 via thiol–maleimide coupling. The labeled substrate sequences for thrombin and factor Xa are abbreviated as s.Thr(sCy5) and s.FXa(sCy5), respectively.…”

Section: Resultsmentioning

confidence: 99%

Bait and Cleave: Exosite-Binding Peptides on Quantum Dots Selectively Accelerate Protease Activity for Sensing with Enhanced Sensitivity

Krause,

Rees,

Darwish

et al. 2024

ACS Nano

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The advantageous optical properties of quantum dots (QDs) motivate their use in a wide variety of applications related to imaging and bioanalysis, including the detection of proteases and their activity. Recent studies have shown that surface chemistry on QDs is able to modulate protease activity, but only nonspecifically. Here, we present a strategy to selectively accelerate the activity of a particular target protease by as much as two orders of magnitude. Exosite-binding "bait" peptides were derived from proteins that span a range of biological roles�substrate, receptor, and inhibitor�and were used to increase the affinity of the QD−peptide conjugates for either thrombin or factor Xa, resulting in increased rates of proteolysis for coconjugated substrates. Unlike effects from QD surface chemistry, the acceleration was specific to the target protease with negligible acceleration of other proteases. Benefits of this "bait and cleave" sensing approach included detection limits that improved by more than an order of magnitude, reenabled detection of target protease against an overwhelming background of nontarget proteolysis, and mitigation of the action of inhibitors. The cumulative results point to a generalizable strategy, where the mechanism of acceleration, considerations for the design of bait peptides and conjugates, and routes to expanding the scope of this approach are discussed. Overall, this research represents a major step forward in the rational design of nanoparticle-based enzyme sensors that enhance sensitivity and selectivity.

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Section: Resultsmentioning

confidence: 99%

Bait and Cleave: Exosite-Binding Peptides on Quantum Dots Selectively Accelerate Protease Activity for Sensing with Enhanced Sensitivity

Krause,

Rees,

Darwish

et al. 2024

ACS Nano

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“…Dye-labeled peptide substrates were conjugated to QDs via the well-established spontaneous binding of polyhistidine tags to the ZnS shell of the QDs. ,, Proteolysis assays were carried out in black polystyrene 384-well microtiter plates using an Infinite M1000 Pro microplate reader (Tecan Ltd., Morrisville, NC). Solutions of QD–peptide conjugate and protease were mixed in equal volumes to a total volume of 40 μL, a final QD concentration of 100 nM, and the desired final concentration of protease.…”

Section: Methodsmentioning

confidence: 99%

Assessing the Steric Impact of Surface Ligands on the Proteolytic Turnover of Quantum Dot–Peptide Conjugates

Krause,

Rees,

Algar

2023

ACS Appl. Mater. Interfaces

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“…Spectroelectrophoresis measurements were made as described previously. 42,43 Additional details for all PL measurements are provided in the ESI. †…”

Section: Pl Measurementsmentioning

confidence: 99%

“…2A). 42,43 PL electropherograms were collected at multiple wavelengths between 530 and 780 nm and the migration time increased as the wavelength increased (Fig. 2B).…”

Section: Characterization By Spectroelectrophoresismentioning

confidence: 99%

Spectrotemporal characterization of photoluminescent silicon nanocrystals and their energy transfer to dyes

et al. 2023

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Preparation and Characterization of Quantum Dot-Peptide Conjugates Based on Polyhistidine Tags

Cited by 3 publications

References 97 publications

Bait and Cleave: Exosite-Binding Peptides on Quantum Dots Selectively Accelerate Protease Activity for Sensing with Enhanced Sensitivity

Bait and Cleave: Exosite-Binding Peptides on Quantum Dots Selectively Accelerate Protease Activity for Sensing with Enhanced Sensitivity

Assessing the Steric Impact of Surface Ligands on the Proteolytic Turnover of Quantum Dot–Peptide Conjugates

Spectrotemporal characterization of photoluminescent silicon nanocrystals and their energy transfer to dyes

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