Serum lipoprotein structure: resonance energy transfer localization of fluorescent lipid probes

Sklar, Larry A.; Doody, Michael C.; Gotto, Antonio M.; Pownall, Henry J.

doi:10.1021/bi00548a005

Cited by 61 publications

(31 citation statements)

References 30 publications

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“…Biophysical studies have demonstrated that these compounds stably insert into phospholipid bilayers with the acyl chain within the hydrophobic interior of the apical leaflet and the hydrophilic fluorescein moiety near the leaflet's phospholipid head groups (Fig. 1b) (Christensen et al, 1999 ;Derzko & Jacobson, 1980 ;Haugland, 1996 ;Sklar et al, 1980 ;Stolz et al, 1992 ;Tocanne et al, 1989). In systems containing double membranes, the probes insert exclusively with the outermost lipid bilayer (Blanco et al, 1994 ;Foley et al, 1986 ;Tocanne et al, 1989).…”

Section: Introductionmentioning

confidence: 99%

Insertion of fluorescent fatty acid probes into the outer membranes of the pathogenic spirochaetes Treponema pallidum and Borrelia burgdorferi

Cox

Radolf

2001

Microbiology

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The authors examined the ability of octadecanoyl (C 18 ), hexadecanoyl (C 16 ) and dodecanoyl (C 12 ) fatty acid (FA) conjugates of 5-aminofluorescein (OAF, HAF and DAF, respectively) to insert into the outer membranes (OMs) of Treponema pallidum, Borrelia burgdorferi and Escherichia coli. Biophysical studies have demonstrated that these compounds stably insert into phospholipid bilayers with the acyl chain within the hydrophobic interior of the apical leaflet and the hydrophilic fluorescein moiety near the phospholipid head groups. Consistent with the known poor intrinsic permeability of the E. coli OM to hydrophobic compounds and surfactants, E. coli was not labelled with any of the FA probes. OAF inserted more readily into OMs of B. burgdorferi than into those of T. pallidum, although both organisms were completely labelled at concentrations at or below 2 µg ml N1 . Intact spirochaetes were labelled with OAF but not with antibodies against known periplasmic antigens, thereby confirming that the probe interacted exclusively with the spirochaetal OMs. Separate experiments in which organisms were cooled to 4 SC (i.e. below the OM phase-transition temperatures) indicated that labelling with OAF was due to insertion of the probe into the OMs. B. burgdorferi, but not T. pallidum, was labelled by relatively high concentrations of HAF and DAF. Taken as a whole, these findings support the prediction that the lack of lipopolysaccharide renders T. pallidum and B. burgdorferi OMs markedly more permeable to lipophilic compounds than their Gram-negative bacterial counterparts. The data also raise the intriguing possibility that these two pathogenic spirochaetes obtain long-chain FAs, nutrients they are unable to synthesize, by direct permeation of their OMs.

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Section: Introductionmentioning

confidence: 99%

Insertion of fluorescent fatty acid probes into the outer membranes of the pathogenic spirochaetes Treponema pallidum and Borrelia burgdorferi

Cox

Radolf

2001

Microbiology

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“…Previous authors have suggested that charged fluorophores associated with lipids should be exposed to the aqueous compartment (Sklar et al, 1980;Arias et al, 1993) and therefore expected to be at the micelle-water interface. We attempted to use quenching by KI as a qualitative verification of the exposure of the fluorophores.…”

Section: Characterization Of Lpsmentioning

confidence: 99%

“…We chose a detergent with low critical micelle concentration (CMC) suitable for fluorescence resonance energy transfer (FRET) (Forster, 1948). FRET has already been shown to provide insight into several aspects of lipid organization including surface density (Fung and Stryer, 1978), placement of fluorescent lipid analogues on or in lipid particles (Sklar et al, 1980), and distances between membranes and protein complexes (Holowka and Baird, 1983;Remmers and Neubig, 1993;Valenzuela et al, 1994). We have used several methods to characterize the interactions of the probe species with the detergent micelles and FRET to define, semiquantitatively, the distances between donor-labeled LPS or lipid donors and lipid acceptors.…”

Section: Introductionmentioning

confidence: 99%

Fluorescence resonance energy transfer analysis of lipopolysaccharide in detergent micelles

Wiström¹,

Jones²,

Tobias³

et al. 1996

Biophysical Journal

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Bacterial endotoxins or lipopolysaccharides (LPS), cell wall components of gram-negative bacteria, are involved in septic shock. LPS consists of a lipid A tail attached to core and O-antigen polysaccharides, but little is known about the supramolecular structure of LPS in blood. We have developed an approach to locate donor and acceptor probes in sulfobetaine palmitate detergent micelles using steady-state and time-resolved fluorescence resonance energy transfer. C18-fluorescein and several LPS species of varying molecular weight labeled with fluorescein isothiocyanate (FITC-LPS) were the donor probes. Acceptor probes were 1,1-dilinoleyl-3,3,3',3'-tetramethyl indocarbocyanine perchlorate (Fast C18-Dil, Ro approximately 68 A), and octadecyl B rhodamine chloride (C18-Rhd, Ro approximately 58 A). With either acceptor, the transfer was of similar high efficiency when FITC-LPS Salmonella minnesota Re 595 (2,500 mol wt, lacking both core and O-antigen) or C18-fluorescein were the fluorescent donor probes. Thus, the donor FITC-LPS with short polysaccharide chain S. minnesota Re 595 and the control donor C18-fluorescein appear to be close to the micelle surface. The transfer efficiency decreased as the molecular weight of the LPS increased. Separation distances between the longest FITC-LPS, S. minnesota (20,000 mol wt, with a long O-antigen), and the micelle were estimated to be 1.5 Ro or more (approximately 100 A), consistent with an extended conformation for the longer O-antigen polysaccharide chain in the detergent.

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“…Other pertinent publications are Baird et al (1979), Doody et al (1983), Eisinger andFlores (1982), Fleming et al (1979), Gutierrez-Merino (1981a, b), Hammes (1981), Holowka and Baird (1983a, b), Issacs et al (1986, Sklar et al (1980), Estep and Thompson (1979), Van der Werf and Ullman (1979). Previous applications of excitation energy transfer in membranes are Cantley and Hammes (1976), Tweet et al (1964), Vanderkooi et al (1977), and Veatch and Stryer (1977).…”

Section: Introductionmentioning

confidence: 99%

Theory for establishing proximity relations in biological membranes by excitation energy transfer measurements

Yguerabide¹

1994

Biophysical Journal

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In a previous publication (Shaklai et al., 1977a) the present author developed a theory for evaluating proximity relations and surface densities sigma in biological membranes by measurements of excitation energy transfer from a donor attached to a specific site of a membrane protein and an acceptor attached to a specific carbon on a membrane lipid. It was assumed that the protein and lipid are randomly distributed in the plane of the membrane and that the donor and acceptor groups are confined to different planes in the membrane separated by a distance Rp. In this article several aspects of the theory presented in the previous paper are clarified, especially noting that the previous theoretical expressions for the time-dependent and steady state fluorescence intensities assumed that the labeled protein molecule is cylindrically symmetric with the symmetry axis perpendicular to the plane of the membrane and that the donor is positioned on the symmetry axis of the protein. This assumption is also implicitly or explicitly made in subsequent formulations by other investigators. In this article we generalize the theory to include the case where the donor is not on the symmetry axis of the labeled protein. Equations for calculating the time-dependent and steady state fluorescence intensities for this more general case are presented, and methods for applying these theoretical expressions to the analysis of steady state fluorescence intensity data and evaluation of proximity parameters are discussed. It is also shown in this article that the linear relation l/lo = 1 + Kq sigma previously derived for simple analysis of excitation transfer data for the condition rc/Ro 1 can be modified to apply to almost all practical ranges of rc/Ro without much affecting its simplicity in the analysis of experimental data.

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Serum lipoprotein structure: resonance energy transfer localization of fluorescent lipid probes

Cited by 61 publications

References 30 publications

Insertion of fluorescent fatty acid probes into the outer membranes of the pathogenic spirochaetes Treponema pallidum and Borrelia burgdorferi

Insertion of fluorescent fatty acid probes into the outer membranes of the pathogenic spirochaetes Treponema pallidum and Borrelia burgdorferi

Fluorescence resonance energy transfer analysis of lipopolysaccharide in detergent micelles

Theory for establishing proximity relations in biological membranes by excitation energy transfer measurements

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