Superficial disposition of the N-terminal region of the surfactant protein SP-C and the absence of specific SP-B–SP-C interactions in phospholipid bilayers

2001 ◽  
Vol 359 (3) ◽  
pp. 651-659 ◽  
Author(s):  
Inés PLASENCIA ◽  
Antonio CRUZ ◽  
Cristina CASALS ◽  
Jesús PÉREZ-GIL
Keyword(s):  
Emission Spectrum ◽  
Protein Interactions ◽  
Fluorescence Emission ◽  
Resonance Energy ◽  
Fold Increase ◽  
Blue Shift ◽  
Terminal Segment ◽  
Phospholipid Bilayers ◽  
Aqueous Environment ◽  
Fluorescence Emission Spectrum

A dansylated form of porcine surfactant-associated protein C (Dns-SP-C), bearing a single dansyl group at its N-terminal end, has been used to characterize the lipid–protein and protein–protein interactions of SP-C reconstituted in phospholipid bilayers, using fluorescence spectroscopy. The fluorescence emission spectrum of Dns-SP-C in phospholipid bilayers is similar to the spectrum of dansyl-phosphatidylethanolamine, and indicates that the N-terminal end of the protein is located at the surface of the membranes and is exposed to the aqueous environment. In membranes containing phosphatidylglycerol (PG), the fluorescence of Dns-SP-C shows a 3-fold increase with respect to the fluorescence of phosphatidylcholine (PC), suggesting that electrostatic lipid–protein interactions induce important effects on the structure and disposition of the N-terminal segment of the protein in these membranes. This effect saturates above 20% PG molar content in the bilayers. The parameters for the interaction of Dns-SP-C with PC or PG have been estimated from the changes induced in the fluorescence emission spectrum of the protein. The protein had similar Kd values for its interaction with the different phospholipids tested, of the order of a few micromolar. Cooling of Dns-SP-C-containing dipalmitoyl PC bilayers to temperatures below the phase transition of the phospholipid produced a progressive blue-shift of the fluorescence emission of the protein. This effect is interpreted as a consequence of the transfer of the N-terminal segment of the protein into less polar environments that originate during protein lateral segregation. This suggests that conformation and interactions of the N-terminal segment of SP-C could be important in regulating the lateral distribution of the protein in surfactant bilayers and monolayers. Potential SP-B–SP-C interactions have been explored by analysing fluorescence resonance energy transfer (RET) from the single tryptophan in porcine SP-B to dansyl in Dns-SP-C. RET has been detected in samples where native SP-B and Dns-SP-C were concurrently reconstituted in PC or PG bilayers. However, the analysis of the dependence of RET on the protein density excluded specific SP-B–Dns-SP-C associations.

A novel ratiometric and reversible fluorescent probe based on naphthalimide for the detection of Al3+ and pH with excellent selectivity

2020 ◽  
Vol 44 (8) ◽  
pp. 3261-3267 ◽  
Author(s):  
Zhuo Li ◽  
Weihong Chen ◽  
Liuyan Dong ◽  
Yan Song ◽  
Ronghang Li ◽  
...  
Keyword(s):  
Emission Spectrum ◽  
Fluorescent Probe ◽  
Fluorescence Probe ◽  
Ph Value ◽  
Fluorescence Emission ◽  
Blue Shift ◽  
Living Cells ◽  
Fluorescence Emission Spectrum ◽  
Excellent Selectivity

A novel ratiometric and reversible fluorescence probe was designed and synthesized. The fluorescent probe recognizes Al3+ to cause a blue shift in the fluorescence emission spectrum. Fluorescence probe can be used as a sensor to detect Al3+ in living cells and Zebra fishes. The probe can detect the pH value when the alkalinity range was 8–10.

Synthesis and solution conformation of [Trp1, Val5]-angiotensin II

1977 ◽  
Vol 55 (1) ◽  
pp. 75-82 ◽  
Author(s):  
Peter W. Schiller
Keyword(s):  
Angiotensin Ii ◽  
Solid Phase ◽  
Fluorescence Emission ◽  
Resonance Energy ◽  
Tryptophan Fluorescence ◽  
Exchange Chromatography ◽  
Phase Method ◽  
Aqueous Environment ◽  
Solution Conformation ◽  
Intramolecular Distance

[Trp1, Val5]-Angiotensin II was synthesized by the solid-phase method and purified by partition chromatography on Sephadex G-25 and by ion-exchange chromatography on Sephadex SP-25. Relative to [Val5]-angiotensin II, the analog displayed 36% smooth muscle activity in a preparation of the superior mesenteric artery. Conformational aspects of the analog were revealed by fluorescence techniques. The fluorescence emission maximum at 350 nm suggests a completely aqueous environment for the tryptophanyl residue in [Trp1, Val5]-angiotensin II. Singlet–singlet resonance energy transfer between Tyr in position 4 and Trp in position 1 was evaluated for a calculation of the intramolecular distance between these two residues on the basis of the Förster equation. From the relative increase of tryptophan fluorescence a transfer efficiency of > 0.9 was obtained, which is compatible with the observed complete quenching of tyrosine fluorescence in the analog. The computed average intramolecular distance of < 8 Å precludes an extended conformation for the N-terminal sequence encompassing residues 1 through 4 at neutral pH and suggests the existence of a loop in this part of the molecule. The results are discussed in relation to the various models proposed for the solution conformation of angiotensin II.

scholarly journals The Synthesis and Application of Nitrogen-Doped Graphene Quantum Dots on Brilliant Blue Detection

2019 ◽  
Vol 2019 ◽  
pp. 1-9 ◽  
Author(s):  
Li Jin ◽  
Ying Wang ◽  
Fengkai Yan ◽  
Jianpo Zhang ◽  
Fangli Zhong
Keyword(s):  
Quantum Dots ◽  
Emission Spectrum ◽  
Graphene Quantum Dots ◽  
Fluorescence Emission ◽  
Fluorescence Decay ◽  
Brilliant Blue ◽  
Fluorescence Emission Spectrum ◽  
Nitrogen Doped ◽  
Nitrogen Doped Graphene ◽  
Doped Graphene

Nitrogen-doped graphene quantum dots had been successfully synthesized and characterized by using transmission electron microscope, X-ray photoelectron spectroscopy, absorbance spectrum, fluorescence emission spectrum, and fluorescence decay curve. TEM results indicated that the diameters of the as-prepared nitrogen-doped graphene quantum dots were in the range of 2 - 5 nm and the lattice space is about 0.276 nm; Raman spectrum result indicated that there were two characteristic peaks, generally named D (~1408 cm−1) and G (~1640 cm−1) bands; both TEM and Raman spectrum results indicated that the as-synthesized product was graphene quantum dots. Deconvoluted high resolution XPS spectra for C1s, O1s, and N1s results indicated that there are -NH-, -COOH, and -OH groups on the surface of nitrogen-doped graphene quantum dot. Fluorescence emission spectrum indicated that the maximum fluorescence emission spectrum of nitrogen-doped graphene quantum dots was blue shift about 30.1 nm and the average fluorescence decay time of nitrogen-doped graphene quantum dots increased about 2 ns, compared with graphene quantum dots without doping of nitrogen. Then, the as-prepared nitrogen-doped graphene quantum dots were used to quantitatively analyze brilliant blue based on the fluorescent quenching of graphene quantum dots, and the effect of pH and reaction time on this fluorescent quenching system was also obtained. Under selected condition, the linear regression equations were F0/F=0.0087 (brilliant blue) + 0.9553 and F0/F=0.01205 (brilliant blue) + 0.6695, and low detection limit was 3.776 μmol/L (3.776 nmol/mL). Once more diluted N-GQDs (0.05 mg/mL) were used, the low detection limit could reach 94.87 nmol/L. Then, temperature-dependent experiment, absorbance spectra, and dynamic fluorescence quenching rate constant were used to study the quenching mechanism; all results indicated that this quenching process was a static quenching process based on the formation of complex between nitrogen-doped graphene quantum dots and brilliant blue through hydrogen bond. Particularly, this method was used to quantitatively analyze the wine sample, of which results have a high consistence with the results of the spectrophotometric method; demonstrating this fluorescence quenching method could be used in practical sample application.

Hydrogen bonded dimers of ketocoumarin in the solid state and alcohol:water binary solvent: fluorescence spectroscopy, crystal structure and DFT investigation

2019 ◽  
Vol 43 (23) ◽  
pp. 9090-9105 ◽  
Author(s):  
Kannan Ramamurthy ◽  
E. J. Padma Malar ◽  
Chellappan Selvaraju
Keyword(s):  
Crystal Structure ◽  
Solid State ◽  
Binary Mixture ◽  
Fluorescence Spectroscopy ◽  
Emission Spectrum ◽  
Fluorescence Emission ◽  
Binary Solvent ◽  
Fluorescence Emission Spectrum ◽  
Hydrogen Bonded

Fluorescence emission spectrum of ketocoumarin dimers in an alcohol:water binary mixture and the solid state.

New insights into the binding behavior of lomefloxacin and human hemoglobin using biophysical techniques: binary and ternary approaches

2019 ◽  
Vol 43 (21) ◽  
pp. 8132-8145 ◽  
Author(s):  
Parisa Mokaberi ◽  
Vida Reyhani ◽  
Zeinab Amiri-Tehranizadeh ◽  
Mohammad Reza Saberi ◽  
Sima Beigoli ◽  
...  
Keyword(s):  
Absorption Spectrum ◽  
Energy Transfer ◽  
Emission Spectrum ◽  
High Probability ◽  
Fluorescence Emission ◽  
Fluorescence Emission Spectrum ◽  
Human Hemoglobin ◽  
Binding Behavior ◽  
Biophysical Techniques

Demonstrates the overlap that had been induced between the fluorescence emission spectrum of Hb and the absorption spectrum of drugs, which has proved that there is a high probability to the occurrence of energy transfer from Hb and LMF in the absence and presence of NRF.

scholarly journals Antagonistic binding of substrates to 3-phosphoglycerate kinase monitored by the fluorescent analogue 2′(3′)-O-(2,4,6-trinitrophenyl)adenosine 5′-triphosphate

1994 ◽  
Vol 301 (3) ◽  
pp. 885-891 ◽  
Author(s):  
M Vas ◽  
A Merli ◽  
G L Rossi
Keyword(s):  
Dissociation Constants ◽  
Equilibrium Dialysis ◽  
Fluorescence Emission ◽  
Phosphoglycerate Kinase ◽  
Structural Data ◽  
Blue Shift ◽  
Competitive Inhibitor ◽  
Intensity Increase ◽  
Fluorescence Emission Spectrum ◽  
Reciprocal Effect

The analogue of ATP, 2′(3′)-O-(2,4,6-trinitrophenyl)adenosine 5′-triphosphate (TNP-ATP), binds tightly to pig muscle 3-phosphoglycerate kinase. A dissociation constant Kd of 0.0095 +/- 0.0015 mM was determined by fluorimetric titration on the basis of 1:1 stoichiometry. TNP-ATP is a strong competitive inhibitor towards MgATP and MgADP with a Ki of 0.008 +/- 0.001 mM for both substrates. It is also a mixed-type inhibitor towards 3-phosphoglycerate with similar inhibition constants. Binding of TNP-ATP to 3-phosphoglycerate kinase is accompanied by a tenfold intensity increase and a blue shift of about 20 nm in its fluorescence emission spectrum and a shift of the pK of its trinitrophenyl group towards a more acidic pH. These findings suggest that the negatively charged trinitrophenyl group of TNP-ATP significantly contributes to the binding of the analogue. By stepwise replacement of the fluorescent TNP-ATP, the dissociation constants (Kd) for ADP and MgADP binding were determined and found to be 0.78 +/- 0.08 and 0.048 +/- 0.006 mM respectively, which are consistent with the values previously determined by equilibrium dialysis [Molnár and Vas (1993) Biochem J. 293, 595-599]. In similar competitive-titration experiments, ATP and MgATP did not completely substitute for TNP-ATP. For the fraction of the analogue that could be substituted, the dissociation constants for MgATP and ATP were estimated to be 0.27 +/- 0.09 and 0.33 +/- 0.15 mM respectively, close to the values determined by equilibrium dialysis. Using the same method, a significant weakening of binding of both (Mg)ADP and (Mg)ATP could be detected in the presence of 3-phosphoglycerate: their respective Kd values became 0.34 +/- 0.04 and 0.51 +/- 0.22 mM. The reciprocal effect, i.e. weakening of 3-phosphoglycerate binding in the presence of the nucleotide substrates, has been observed previously [Vas and Batke (1984) Eur. J. Biochem. 139, 115-123]. Similarly, a much weaker binding of (Mg)ATP could be observed in the presence of 1,3-bisphosphoglycerate (Kd = 2.30 +/- 0.68 mM). The possible reason for the mutual weakening of substrate binding is discussed in the light of the available structural data.

scholarly journals A quantitative protocol for dynamic measurements of protein interactions by Förster resonance energy transfer-sensitized fluorescence emission

2008 ◽  
Vol 6 (suppl_1) ◽  
Author(s):  
A.D Elder ◽  
A Domin ◽  
G.S Kaminski Schierle ◽  
C Lindon ◽  
J Pines ◽  
...  
Keyword(s):  
Energy Transfer ◽  
Protein Interactions ◽  
Imaging Techniques ◽  
Resonance Energy Transfer ◽  
Fluorescence Emission ◽  
Resonance Energy ◽  
Forster Resonance Energy Transfer ◽  
Confocal Imaging ◽  
Förster Resonance Energy Transfer ◽  
Förster Resonance

Fluorescence detection of acceptor molecules sensitized by Förster resonance energy transfer (FRET) is a powerful method to study protein interactions in living cells. The method requires correction for donor spectral bleed-through and acceptor cross-excitation as well as the correct normalization of signals to account for varying fluorophore concentrations and imaging parameters. In this paper, we review different methods for FRET signal normalization and then present a rigorous model for sensitized emission measurements, which is both intuitive to understand and practical to apply. The method is validated by comparison with the acceptor photobleaching and donor lifetime-imaging techniques in live cell samples containing EYFP and ECFP tandem constructs exhibiting known amounts of FRET. By varying the stoichiometry of interaction in a controlled fashion, we show that information on the fractions of interacting donors and acceptors can be recovered. Furthermore, the method is tested by performing measurements on different microscopy platforms in both widefield and confocal imaging modes to show that signals recovered under different imaging conditions are in quantitative agreement. Finally, the method is applied in the study of dynamic interactions in the cyclin–cdk family of proteins in live cells. By normalizing the obtained signals for both acceptor and donor concentrations and using a FRET exhibiting control construct for calibration, stoichiometric changes in these interactions could be visualized in real time. The paper is written to be of practical use to researchers interested in performing sensitized emission measurements. The correct interpretation of the retrieved signals in a biological context is emphasized, and guidelines are given for the practical application of the developed algorithms.

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