Highly Fluorescent Quantum Dots through Protein Design

Due to their high emission quantum yields, excellent resistance to photobleaching, photostability, and large Stokes shifts, compared to conventional organic fluorescent dyes, Quantum dots (QDs) have gained a great deal of attention. Lately, significant efforts have been made to use QDs, for example, for bioimaging, as fluorescent reporters for biomedical applications. However the synthesis of traditional chemical QDs generally includes toxic reagents, organic solvents, intense reaction conditions, intensive energy input, and relies on a variety of tedious and time-consuming steps. Additionally, QDs require postsynthetic processing after chemical synthesis to make them water-soluble for biological applications. Lastly, most QDs synthesised by chemical methods show low biocompatibility and poor stability under conditions of high ionic strength, thereby significantly limiting their use.

This research was carried out at Center for Cooperative Research in Biomaterials (CIC biomaGUNE), Basque Research and Technology Alliance and publish in chemistry of Materials under title " Protein Design for the Synthesis and Stabilization of Highly Fluorescent Quantum Dots"

This work validates protein engineering as a new approach to eco-friendly and sustainable aqueous routes for the sustainable synthesis and stabilisation of high-quality fluorescent QDs. The focus of this research was CdS QDs, as this composition has been extensively studied, but this approach may be easily converted to QDs for other compositions of metal. The inclusion of unique metal coordination sites that depend on histidines and cysteines has established a flexible approach to the design of proteins for the green synthesis of highly fluorescent CdS QDs. 

Protein architecture emerges here as a promising method in various applications such as light-emitting devices, metal ion detection, and biomedical applications to produce protein-hybrid nanomaterials as widely applicable instruments.

For the sustainable synthesis and stabilisation of biocompatible CdS QDs, this study introduces a protein engineering approach to integrating metal coordination sites. The resulting protein-stabilized CdS QDs (Prot-QDs), generated at 37 C by a green aqueous path, are highly photoluminescent and photostable, have a long shelf life and under physiological conditions exhibit high stability. The Prot-QDs showed productive results

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The authors observe excellent Internalization and high cell fluorescence, also at low and biocompatible dots. They further explain " as this structure has been extensively studied, but this method could easily be translated with other metal compositions into QDs". 

Four cysteines were inserted into the fluorescent properties of the Prot-QDs in order to research the effect of the metal coordinating protein residue on the fluorescent properties of the Prot-QDs and to explore the tunability of the QD properties of the protein template. Site of  generating the C module. for the synthesis and stabilisation of CdS QDs, Wt(C)4Wt protein was assessed. The fluorescence emission of the cysteine stabilised CdS QDs is lower than the Wt(H)4 Wt stabilised Prot-QDs. The QY of the Prot-QDs was therefore reduced (QY = 24 percent) compared to the previously mentioned (QY = 32 percent) for histidine coordination when cysteines are the metal coordinating residues.

The metal/protein ratio's effect (from 20:1 to 80:1) was evaluated for the the synthesis and stabilisation of CdS QDs was evaluated by using (Wt)8-histag and Wt (H)4 Wt. At an 80:1 of metals/proteins ratio, the highest fluorescence emission intensity of the Prot-QDs was observed.

The protein aggregation was greater than 80:1 and the fluorescence emission intensity subsequently decreased. Furthermore, using the Wt(H)4 Wt protein, the effect of the Cd/Na2S ratio (from 1:0.4 to 1:1.6) on the fluorescent properties of the resulting Prot-QDs was evaluated. The largest Wt(H)4 Wt-CdS fluorescence emission strength was observed at a ratio of 1:0.8. A red change in the Wt(H)4Wt-CdS fluorescence emission was also observed when the Cd/Na2S ratio decreased from 1:0.4 to 1:1.6, which could be correlated with an increase in the size of nanocrystals.

In contrast to the poly histidine fusion protein and GSH, engineered protein templates not only improve the PL properties of Prot-QDs, but also improve their photostability, stability under physiological simulated conditions, and biocompatibility. The resulting engineered Prot-QDs are highly photoluminescent and photostable, created by a green aqueous route at 37° C.

They have a long shelf life, and stable under simulated physiological conditions, showing high biocompatibility and low manufacturing costs. Finally, without affecting cell viability, Prot-QDs are able to enter living cells, showing great cell labelling potential at very low doses, making them useful tools for biomedical applications. In addition, the method built in this study could be translated into other architectures of the protein cage (presenting structures of alpha-helices), which were previously used as models for the synthesis of various nanomaterials, in order to adapt the properties of the resulting protein-hybrid nanomaterial to different fields for their application.