There remains a gap in understanding the enzyme interactions with the coacervate as a substrate hub. Here, we study how the hydrophobicity nature of coacervate affects the interactions of the embedded substrate with a protease. We design oligopeptide-based coacervates that comprise an anionic Asp-peptide (D10) and a cationic Arg-peptide (R5R5) with a proteolytic cleavage site. The coacervates dissolve when exposed to the main protease. We exploit the condensed structure, implement a self-quenching mechanism, and characterize enzyme kinetics with Cy5.5-labeled peptides. The determined specificity constant is 5,817 M-1 s-1 and is similar to that of the free substrate. We further show that the enzyme kinetics depend on the amount of dye incorporated into the coacervates. Our work presents a simple design for coacervates with tuned bioactivities and provides insights into the kinetics between the enzyme and coacervates as a substrate hub.
We report on the use of lipoic acid-appended polyethylene glycol (LA-PEG)-stabilized AuNPs, covalently coupled to dye-labelled peptides, as sensitive optically-addressable sensors for determining the catalytic efficiency of the human matrix metalloproteinase-14 (MMP-14), a cancer biomarker. We exploit real-time dye PL recovery triggered by MMP-14 hydrolysis of the AuNP-tethered peptide-dye to extract quantitative analysis of the proteolysis kinetics. Sub-nanomolar limit of detections for MMP-14 has been achieved using our hybrid bioconjugates.
Significance: Photoacoustic imaging has shown advantages over the periodontal probing method in measuring the periodontal probing depth, but the large size of conventional photoacoustic transducers prevents imaging of the more posterior teeth.
Aim: Our aim is to develop a photoacoustic imaging system to image the more posterior periodontal pocket.
Approach: We report a clinical “hockey-stick”-style transducer integrated with fibers for periodontal photoacoustic imaging. Cuttlefish ink labeled the periodontal pocket as the photoacoustic contrast agent.
Results: We characterized the imaging system and then measured the pocket depth of 35 swine teeth. Three raters evaluated the performance of the hockey-stick transducer. The measurements between the Williams probing (gold standard) and the photoacoustic methods were blinded but highly correlated. We showed a bias of ∼0.3 mm for the imaging-based technique versus Williams probing. The minimum inter-reliability was over 0.60 for three different raters of varying experience, suggesting that this approach to measure the periodontal pocket is reproducible. Finally, we imaged three pre-molars of a human subject. We could access more upper and posterior teeth than conventional linear transducers.
Conclusions: The unique angle shape of the hockey-stick transducer allows it to image more posterior teeth than regular linear transducers. This study demonstrated the ability of a hockey-stick transducer to measure the periodontal pocket via photoacoustic imaging.
N-heterocyclic carbenes (NHCs) have attracted tremendous attention over the past decade, as it is expected to form strong coordination to transition metal complexes and surfaces. Here, we investigate the interactions between colloidal gold nanoparticles (AuNPs), or luminescent quantum dots (QDs) and a multidentate NHC-based polymer ligand. The ligand design relies on the nucleophilic addition reaction between several NHC anchoring groups, short polyethylene glycol (PEG) blocks, and a polymer chain. We find that such NHC-decorated ligands rapidly coordinate onto both sets of nanocrystals, which is attributed to the inherent σ-donating nature (soft Lewis base) of NHC groups combined with the soft Lewis acidic character of nanocrystal surfaces. We combine NMR spectroscopy, fluorescence spectroscopy, high-resolution transmission electron microscopy and dynamic light scattering to characterize the NHCstabilized nanocrystals and gain insights into the nature of the binding interactions. In particular, we find that the newly coated nanocrystals exhibit long-term colloidal stability over a broad range of conditions with no sign of degradation or aggregation build up, while preserving their photophysical properties, for at least one year of storage.
Coating inorganic nanocrystals (e.g., quantum dots and gold nanoparticles) with polymer ligands presenting many lipoic acid (LA) anchoring groups provides them with excellent colloidal stability in aqueous media. Here we exploit the natural swelling of polymer macromolecules, which imposes a configuration that leaves a fraction of the anchors on the polymerstabilized nanocolloids free or uncoordinated and target them for conjugation using thiol-to-maleimide chemistry. This allows easy surface functionalization of the nanocrystals, without the need to introduce additional reactive groups. We apply a photoligation strategy to coat QDs and AuNPs, followed by coupling with maleimide-modified dyes. We then use optical absorption and resonance energy transfer measurements, to extract estimates for the fraction of accessible LAs per nanocrystal. To further prove the effectiveness of this approach, we construct a ratiometric pH sensing probe made of QDSNARF conjugates. The combination of the multi-coordinating ligand design and in-situ photoligation yields colloidally stable nanocrystals, presenting several thiol reactive sites. Our results are promising and could advance the integration of nanomaterials in biological sensing and imaging applications.
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