Abstract
An M13 bacteriophage-based Förster resonance energy transfer (FRET) system is developed to estimate intermolecular distance at the nanoscale using a complex of CdSSe/ZnS nanocrystal quantum dots, genetically engineered M13 bacteriophages labeled with fluorescein isothiocyanate and trinitrotoluene (TNT) as an inhibitor. In the absence of trinitrotoluene, it is observed that a significant spectral shift from blue to green occur, which represents efficient energy transfer through dipole-dipole coupling between donor and acceptor, or FRET-on mode. On the other hand, in the presence of trinitrotoluene, the energy transfer is suppressed, since the donor-to-acceptor intermolecular distance is detuned by the specific capturing of TNT by the M13 bacteriophage, denoted as FRET-off mode. These noble features are confirmed by changes in the fluorescence intensity and the fluorescence decay curve. TNT addition to our system results in reducing the total energy transfer efficiency considerably from 16.1% to 7.6% compared to that in the non-TNT condition, while the exciton decay rate is significantly enhanced. In particular, we confirm that the energy transfer efficiency satisfies the original intermolecular distance dependence of FRET. The relative donor-to-acceptor distance is changed from 70.03 Å to 80.61 Å by inclusion of TNT.
Highlights
In the biological recognition process, the molecular interaction between receptors and analytes is often associated with a conformational change due to specific physical or chemical binding[1,2,3]
We designed an M13 phage-based Förster resonance energy transfer (FRET) system using a complex of water-soluble CdSSe/ZnS nanocrystal quantum dots, a genetically engineered M13 bacteriophage labeled with fluorescein isothiocyanate and trinitrotoluene (TNT) as an inhibitor
nanocrystal quantum dots (NQDs) were positively charged by a polydiallydimethyl-ammounium chloride (PDDA) organic coating layer and had no linkable functional groups
Summary
In the biological recognition process, the molecular interaction between receptors and analytes is often associated with a conformational change due to specific physical or chemical binding[1,2,3]. FRET-based sensing facilitates the visualization of receptor-analyte interactions through the detection of color change and provides clues regarding relative intermolecular distances between reacting molecules through time-integrated or time-resolved analysis. Considering that resonant coupling between dipoles is within 100 Å60, the dimensions of the M13 phage are much larger On this account, the M13 phage is often used in a single-molecular FRET scheme, whereby a donor and acceptor pair is immobilized onto the M13 phage[56,57,58]. The intermolecular distance between them is about 24~32 Å56 This restriction can affect the sensitivity of FRET-based analyte sensing because it limits the number of specific peptides of the M13 phage eligible to participate in receptor-analyte reactions. We estimated the relative intermolecular distance between a donor and acceptor based on the energy transfer efficiency
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