Using a Novel Dual Fluorescence Quenching Assay for Measurement of Tryptophan Depth within Lipid Bilayers To Determine Hydrophobic α-Helix Locations within Membranes†

Biochemistry ◽  
2003 ◽  
Vol 42 (11) ◽  
pp. 3265-3274 ◽  
Author(s):  
Gregory A. Caputo ◽  
Erwin London
Keyword(s):  
Fluorescence Quenching ◽  
Lipid Bilayers ◽  
Dual Fluorescence ◽  
Α Helix ◽  
Quenching Assay

The Inhibition of Polysialyltranseferase ST8SiaIV Through Heparin Binding to Polysialyltransferase Domain (PSTD)

2019 ◽  
Vol 15 (5) ◽  
pp. 486-495 ◽  
Author(s):  
Li-Xin Peng ◽  
Xue-Hui Liu ◽  
Bo Lu ◽  
Si-Ming Liao ◽  
Feng Zhou ◽  
...  
Keyword(s):  
Fluorescence Quenching ◽  
Polysialic Acid ◽  
Neuronal Cell ◽  
Cd Spectroscopy ◽  
Migration And Invasion ◽  
Heparin Binding ◽  
Clinical Prognosis ◽  
Quenching Analysis ◽  
Different Types ◽  
Α Helix

Background:The polysialic acid (polySia) is a unique carbohydrate polymer produced on the surface Of Neuronal Cell Adhesion Molecule (NCAM) in a number of cancer cells, and strongly correlates with the migration and invasion of tumor cells and with aggressive, metastatic disease and poor clinical prognosis in the clinic. Its synthesis is catalyzed by two polysialyltransferases (polySTs), ST8SiaIV (PST) and ST8SiaII (STX). Selective inhibition of polySTs, therefore, presents a therapeutic opportunity to inhibit tumor invasion and metastasis due to NCAM polysialylation. Heparin has been found to be effective in inhibiting the ST8Sia IV activity, but no clear molecular rationale. It has been found that polysialyltransferase domain (PSTD) in polyST plays a significant role in influencing polyST activity, and thus it is critical for NCAM polysialylation based on the previous studies.Objective:To determine whether the three different types of heparin (unfractionated hepain (UFH), low molecular heparin (LMWH) and heparin tetrasaccharide (DP4)) is bound to the PSTD; and if so, what are the critical residues of the PSTD for these binding complexes?Methods:Fluorescence quenching analysis, the Circular Dichroism (CD) spectroscopy, and NMR spectroscopy were used to determine and analyze interactions of PSTD-UFH, PSTD-LMWH, and PSTD-DP4.Results:The fluorescence quenching analysis indicates that the PSTD-UFH binding is the strongest and the PSTD-DP4 binding is the weakest among these three types of the binding; the CD spectra showed that mainly the PSTD-heparin interactions caused a reduction in signal intensity but not marked decrease in α-helix content; the NMR data of the PSTD-DP4 and the PSTDLMWH interactions showed that the different types of heparin shared 12 common binding sites at N247, V251, R252, T253, S257, R265, Y267, W268, L269, V273, I275, and K276, which were mainly distributed in the long α-helix of the PSTD and the short 3-residue loop of the C-terminal PSTD. In addition, three residues K246, K250 and A254 were bound to the LMWH, but not to DP4. This suggests that the PSTD-LMWH binding is stronger than the PSTD-DP4 binding, and the LMWH is a more effective inhibitor than DP4.Conclusion:The findings in the present study demonstrate that PSTD domain is a potential target of heparin and may provide new insights into the molecular rationale of heparin-inhibiting NCAM polysialylation.

Mathematical simulation of dual fluorescence quenching

2014 ◽  
Vol 116 (3) ◽  
pp. 360-364 ◽  
Author(s):  
V. A. Morozov ◽  
N. D. Chuvylkin ◽  
E. A. Smolenskii
Keyword(s):  
Fluorescence Quenching ◽  
Mathematical Simulation ◽  
Dual Fluorescence

Amine-rich carbon nitride nanoparticles: Synthesis, covalent functionalization with proteins and application in a fluorescence quenching assay

Nano Research ◽  
2019 ◽  
Vol 12 (8) ◽  
pp. 1862-1870 ◽  
Author(s):  
Gabriele Capilli ◽  
Simone Cavalera ◽  
Laura Anfossi ◽  
Cristina Giovannoli ◽  
Marco Minella ◽  
...  
Keyword(s):  
Fluorescence Quenching ◽  
Carbon Nitride ◽  
Covalent Functionalization ◽  
Nanoparticles Synthesis ◽  
Quenching Assay

Different Effects of Di- and Triphenyltin Compounds on Lipid Bilayer Dithionite Permeabilization

2000 ◽  
Vol 55 (9-10) ◽  
pp. 758-763 ◽  
Author(s):  
Janina Gabrielska ◽  
Teresa Kral ◽  
Marek Langner ◽  
Stanislaw Przestalski
Keyword(s):  
Molecular Structure ◽  
Lipid Bilayer ◽  
Lipid Bilayers ◽  
Membrane Properties ◽  
Human Environment ◽  
Vesicle Size ◽  
Phenyl Rings ◽  
Positively Charged ◽  
Steric Constraints ◽  
Quenching Assay

Abstract Phenyltins are chemicals widely used in industry, hence their occurrence in the human environment is frequent and widespread. Such compounds include hydrophobic phenyl rings bonded to positively charged tin. This molecular structure makes them capable of adsorbing onto and penetrating through biological membranes, hence they are potentially hazardous. Two such compounds, diphenyltin and triphenyltin, show different steric constraints when interacting with the lipid bilayer. It has been demonstrated that these compounds are positioned at different locations within model lipid bilayers, causing dissimilarity in their ability to affect membrane properties. In this paper we present a study regarding the ability of these two phenyltins to facilitate the transport of S2O4-2 ions across the lipid bilayer, evaluated by a fluorescence quenching assay. In concentration range of up-to 60 μm those compounds do not affect lipid bilayer topology, when evaluated by vesicle size distribution. Both phenyltins facilitate the transfer of S2O4-2 across the model lipid bilayer, but the dependence of dithionite transport on phenyltin concentration is different for both. In principle, above 20 μm triphenyltin is more efficient in transfering ions across the lipid bilayer than diphenyltin.

Viral Fusion Peptides: A Tool Set to Disrupt and Connect Biological Membranes

2000 ◽  
Vol 20 (6) ◽  
pp. 501-518 ◽  
Author(s):  
Lukas K. Tamm ◽  
Xing Han
Keyword(s):  
Lipid Bilayers ◽  
Biological Membrane ◽  
Membrane Curvature ◽  
Current Evidence ◽  
Viral Fusion ◽  
Fusion Peptides ◽  
Membrane Surfaces ◽  
Self Association ◽  
Α Helix ◽  
And Function

The structure and function of viral fusion peptides are reviewed. The fusion peptides of influenza virus hemagglutinin and human immunodeficiency virus are used as paradigms. Fusion peptides associated with lipid bilayers are conformationally polymorphic. Current evidence suggests that the fusion-promoting state is the obliquely inserted α-helix. Fusion peptides also have a tendency to self-associate into γ-sheets at membrane surfaces. Although the conformational conversion between α- and γ-states is reversible under controlled conditions, its physiological relevance is not yet known. The energetics of peptide insertion and self-association could be measured recently using more soluble “second generation” fusion peptides. Fusion peptides have been reported to change membrane curvature and the state of hydration of membrane surfaces. The combined results are built into a model for the mechanism by which fusion peptides are proposed to assist in biological membrane fusion.

scholarly journals Bactericidal and membrane disruption activities of the eosinophil cationic protein are largely retained in an N-terminal fragment

2009 ◽  
Vol 421 (3) ◽  
pp. 425-434 ◽  
Author(s):  
Marc Torrent ◽  
Beatriz G. de la Torre ◽  
Victòria M. Nogués ◽  
David Andreu ◽  
Ester Boix
Keyword(s):  
Lipid Bilayers ◽  
Cell Damage ◽  
Eosinophil Cationic Protein ◽  
Lipid Vesicles ◽  
Antimicrobial Activities ◽  
Membrane Disruption ◽  
Structural Determinants ◽  
Cationic Protein ◽  
Native Proteins ◽  
Α Helix

ECP (eosinophil cationic protein) is an eosinophil secretion protein with antipathogen activities involved in the host immune defence system. The bactericidal capacity of ECP relies on its action on both the plasma membrane and the bacterial wall. In a search for the structural determinants of ECP antimicrobial activity, we have identified an N-terminal domain (residues 1–45) that retains most of ECP's membrane-destabilizing and antimicrobial activities. Two sections of this domain, ECP-(1–19) and ECP-(24–45), have also been evaluated. All three peptides bind and partially insert into lipid bilayers, inducing aggregation of lipid vesicles and leakage of their aqueous content. In such an environment, the peptides undergo conformational change, significantly increasing their α-helix content. The bactericidal activity of the three peptides against Escherichia coli and Staphylococcus aureus has been assessed at both the cytoplasmic membrane and the bacterial envelope levels. ECP-(1–45) and ECP-(24–45) partially retain the native proteins ability to bind LPS (lipopolysaccharides), and electron microscopy reveals cell damage by both peptides. Interestingly, in the E. coli cells agglutination activity of ECP is only retained by the longest segment ECP-(1–45). Comparative results suggest a task distribution, whereby residues 1–19 would contribute to membrane association and destabilization, while the 24–45 region would be essential for bactericidal action. Results also indicate that ECP cytotoxicity is not uniquely dependant on its membrane disruption capacity, and that specific interactions at the bacteria wall are also involved.

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