Di-Arginine Additives for Dissociation of Gold Nanoparticle Aggregates: A Matrix-Insensitive Approach with Applications in Protease Detection

Abstract

We report the reversible aggregation of gold nanoparticles (AuNPs) assemblies via a di-arginine peptide additive and thiolated PEGs (HS-PEGs). The AuNPs were first aggregated by attractive forces between the citrate-capped surface and the arginine side chains. We found that the HS-PEG thiol group has a higher affinity for the AuNP surface, thus leading to redispersion and colloidal stability. In turn, there was a robust and obvious color change due to on/off plasmonic coupling. The assemblies' dissociation was directly related to the HS-PEG structural properties such as their size or charge. As an example, HS-PEGs with a molecular weight below 1 kDa could dissociate 100% of the assemblies and restore the exact optical properties of the initial AuNP suspension (prior to the assembly). Surprisingly, the dissociation capacity of HS-PEGs was not affected by the composition of the operating medium and could be performed in complex matrices such as plasma, saliva, bile, urine, cell lysates, or even seawater. The high affinity of thiols for the gold surface encompasses by far the one of endogenous molecules and is thus favored. Moreover, starting with AuNPs already aggregated ensured the absence of a background signal as the dissociation of the assemblies was far from spontaneous. Remarkably, it was possible to dry the AuNP assemblies and solubilize them back with HS-PEGs, improving the colorimetric signal generation. We used this system for protease sensing in biological fluids. Trypsin was chosen as the model enzyme, and highly positively charged peptides were conjugated to HS-PEG molecules as cleavage substrates. The increase of positive charge of the HS-PEG−peptide conjugate quenched the dissociation capacity of the HS-PEG molecules, which could only be restored by the proteolytic cleavage. Picomolar limit of detection was obtained as well as the detection in saliva or urine.

A Protease-Responsive Polymer/Peptide Conjugate and Reversible Assembly of Silver Clusters for the Detection of Porphyromonas gingivalis Enzymatic Activity

“…This can be explained by the higher affinity of (RRK) 2 and (RRK) 3 for the citrate layer compared to RRK. Indeed, the reversible assembly works only if the citrate/peptide layer can be removed from the surface by the stabilizing ligand …”

Empirical Optimization of Peptide Sequence and Nanoparticle Colloidal Stability: The Impact of Surface Ligands and Implications for Colorimetric Sensing

“…The aggregation of metallic colloids leads to a bathochromic shift in their surface plasmon resonance (SPR) band and results in a pronounced color change. , The ultimate color formation is a function of the core composition, particle morphology, − and surface chemistry, , which modulate the resonant coupling of light and free electrons in metallic nanostructures. For instance, the aggregation of gold nanoparticles (AuNPs) dramatically changes the dispersion color visually from red to purple/blue by the naked eye due to the strong sensitivity of SPR to the interparticle distance combined with the high molar absorption coefficients. , Such aggregation-induced plasmonic coupling has been exploited as an optical signal transduction strategy in colorimetric sensors with widespread use in bioanalytical applications. − …”

“…). Over the past two years, our group has designed a set of colorimetric sensors to study proteases through an array of NP types, proteases, and aggregation mechanisms. ,,,− However, we have not yet systematically investigated the effects of surface chemistry. Surface chemistry plays a critical role in endowing colloidal stability and interfacial functionalitythese factors determine how nanoparticles adapt to the chemical or environmental stimuli and thus manifest in interparticle crosslinking and colorimetric sensing. ,− The depth and diversity of the ligand field provide choices spanning from metal complexes, small organic compounds, , polymers, , and other biomacromolecules ( e.g.…”

Goldilocks Energy Minimum: Peptide-Based Reversible Aggregation and Biosensing

“…Although a few studies have reported aggrega-T h i s c o n t e n t i s tion-based colorimetric assays in biofluids, they often require sample dilution or removal of the analyte from the matrix. 6,14,15 Finally, AuNP aggregation can amplify eventually leading to aggregates larger than 1 μm that lose colloidal stability and become colorless. 3 To address these issues, we recently proposed colorimetric assays based on dissociation.…”

“…3 To address these issues, we recently proposed colorimetric assays based on dissociation. 14 Here, particle aggregation was first triggered by a short cationic peptide containing arginine and lysine that induced plasmonic coupling of anionic AuNPs coated with citrate. Most importantly, this aggregation was reversible upon addition of steric stabilizers such as thiol-PEG molecules (HS-PEGs).…”