Use of Fluorescence Resonance Energy Transfer for Rapid Detection of Enteroviral Infection In Vivo

Hwang, Yu-Chen; Chen, Wilfred; Yates, Marylynn V.

doi:10.1128/aem.72.5.3710-3715.2006

Cited by 37 publications

(32 citation statements)

References 48 publications

(18 reference statements)

Supporting

Mentioning

Contrasting

Order By: Relevance

“…We next utilized a bacterial selection to identify cyclic peptides that inhibit a protease of interest. Several such schemes have been described in the literature for protease inhibitors (25)(26)(27)(28)(29)(30)(31)(32)(33)(34)(35) in which the viability of E. coli is linked to protease activity (32,33). For example, Block and Grafstrom (32) introduced an HIV protease recognition sequence into the tetracycline resistance gene (Tn10), a metal-tetracycline/ H þ antiporter that contains two functional, homologous integral membrane domains (TetA and TetB) linked by a central cytosolic hinge (36).…”

Section: Resultsmentioning

confidence: 99%

Evolution of cyclic peptide protease inhibitors

Young

Ahmad

et al. 2011

Proc. Natl. Acad. Sci. U.S.A.

120

127

View full text Add to dashboard Cite

We report a bacterial system for the evolution of cyclic peptides that makes use of an expanded set of amino acid building blocks. Orthogonal aminoacyl-tRNA synthetase/tRNA CUA pairs, together with a split intein system were used to biosynthesize a library of ribosomal peptides containing amino acids with unique structures and reactivities. This peptide library was subsequently used to evolve an inhibitor of HIV protease using a selection based on cellular viability. Two of three cyclic peptides isolated after two rounds of selection contained the keto amino acid p -benzoylphenylalanine ( p BzF). The most potent peptide (G12: GIXVSL; X = p BzF) inhibited HIV protease through the formation of a covalent Schiff base adduct of the p BzF residue with the ϵ-amino group of Lys 14 on the protease. This result suggests that an expanded genetic code can confer an evolutionary advantage in response to selective pressure. Moreover, the combination of natural evolutionary processes with chemically biased building blocks provides another strategy for the generation of biologically active peptides using microbial systems.

show abstract

Section: Resultsmentioning

confidence: 99%

Evolution of cyclic peptide protease inhibitors

Young

Ahmad

et al. 2011

Proc. Natl. Acad. Sci. U.S.A.

120

127

View full text Add to dashboard Cite

show abstract

“…This reporter assay was performed as described previously (4). Briefly, cells were cultured in 16-well slide chambers to confluence, and monolayers containing approximately 10 5 cells were infected with viruses as indicated below.…”

mentioning

confidence: 99%

Comparison of a Reporter Assay and Immunomagnetic Separation Real-Time Reverse Transcription-PCR for the Detection of Enteroviruses in Seeded Environmental Water Samples

Hwang¹,

Leong²,

Chen

et al. 2007

Appl Environ Microbiol

Self Cite

View full text Add to dashboard Cite

Two newly developed protocols for infective virus detection were compared to the plaque assay. An immunomagnetic separation procedure coupled with real-time reverse transcription-PCR of viral nucleic acids was developed to identify intact enteroviral particles, and a reporter cell system responding to viral replication based on fluorescent resonance energy transfer for detection of infectious enteroviruses was tested. Both new procedures detected infective viruses in environmental samples at the same level as the plaque assay.

show abstract

“…Plus, when a fusion protein consisting of calmodulin and the M13 peptide was flanked by two fluorescent protein variants, the calcium binding to the calmodulin moiety and the consequent affinity to the M13 peptide were found to result in a stronger FRET intensity between the fluorescent proteins (19). Thus, the protein and the improved derivatives have been applied to monitor the calcium concentrations in living cells (1,13), while various FRET-based sensors have been developed for the detection of cyclic AMP (29), cGMP (11,24), GTP/GDP (20), inositol 1,4,5-trisphosphate (25,27), and infectious enteroviruses (12).…”

mentioning

confidence: 99%

Design and Application of Highly Responsive Fluorescence Resonance Energy Transfer Biosensors for Detection of Sugar in LivingSaccharomyces cerevisiaeCells

Song

Lee

et al. 2007

Appl Environ Microbiol

View full text Add to dashboard Cite

A protein sensor with a highly responsive fluorescence resonance energy transfer (FRET) signal for sensing sugars in living Saccharomyces cerevisiae cells was developed by combinatorial engineering of the domain linker and the binding protein moiety. Although FRET sensors based on microbial binding proteins have previously been created for visualizing various sugars in vivo, such sensors are limited due to a weak signal intensity and a narrow dynamic range. In the present study, the length and composition of the linker moiety of a FRET-based sensor consisting of CFP-linker 1 -maltose-binding protein-linker 2 -YFP were redesigned, which resulted in a 10-fold-higher signal intensity. Molecular modeling of the composite linker moieties, including the connecting peptide and terminal regions of the flanking proteins, suggested that an ordered helical structure was preferable for tighter coupling of the conformational change of the binding proteins to the FRET response. When the binding site residue Trp62 of the maltose-binding protein was diversified by saturation mutagenesis, the Leu mutant exhibited an increased binding constant (82 M) accompanied by further improvement in the signal intensity. Finally, the maltose sensor with optimized linkers was redesigned to create a sugar sensor with a new specificity and a wide dynamic range. When the optimized maltose sensors were employed as in vivo sensors, highly responsive FRET images were generated from real-time analysis of maltose uptake of Saccharomyces cerevisiae (baker's yeast).Fluorescence resonance energy transfer (FRET) is a nonradiative energy transfer between donor and acceptor fluorophores in which the signal intensity changes depending on the proximity and relative angular orientation between the fluorophores (15). As such, FRETs between fluorescent proteins with a spectral overlap have been used to investigate spatial and temporal interactions in living cells when they are conjugated to interacting protein pairs (9,18,28). Plus, when a fusion protein consisting of calmodulin and the M13 peptide was flanked by two fluorescent protein variants, the calcium binding to the calmodulin moiety and the consequent affinity to the M13 peptide were found to result in a stronger FRET intensity between the fluorescent proteins (19). Thus, the protein and the improved derivatives have been applied to monitor the calcium concentrations in living cells (1, 13), while various FRET-based sensors have been developed for the detection of cyclic AMP (29), cGMP (11, 24), GTP/GDP (20), inositol 1,4,5-trisphosphate (25, 27), and infectious enteroviruses (12).The hinge-like twisting and bending motion of bacterial periplasmic binding proteins has also been introduced as an element for FRET-based sensors (7,8). Here, the substrate is located deep within the cleft between the globular N and C lobes, and in the case of substrate binding, the protein undergoes a hinge-like bending motion, causing a change in distance and/or angular orientation between the fluorescent proteins. Maltose-...

show abstract

Use of Fluorescence Resonance Energy Transfer for Rapid Detection of Enteroviral Infection In Vivo

Cited by 37 publications

References 48 publications

Evolution of cyclic peptide protease inhibitors

Evolution of cyclic peptide protease inhibitors

Comparison of a Reporter Assay and Immunomagnetic Separation Real-Time Reverse Transcription-PCR for the Detection of Enteroviruses in Seeded Environmental Water Samples

Design and Application of Highly Responsive Fluorescence Resonance Energy Transfer Biosensors for Detection of Sugar in LivingSaccharomyces cerevisiaeCells

Contact Info

Product

Resources

About