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ABSTRACT: It has been reported that the oxidation of phosphatidylcholine (PC) is necessary for C-reactive protein (CRP) to bind to lipid membranes, but it remains elusive why CRP only binds oxidized membranes. Here we offer a new perspective on the role of membrane curvature in CRP binding using engineered lipoprotein particle (LPP) mimics. We show that CRP binds preferentially to LPP mimics with diameters of #28 nm, and binding of CRP to these mimics leads to the dissociation of native CRP into monomeric CRP, exposing CRP neo-epitopes that bind C1q. We also show that the smaller LPP mimics compete for CRP binding to oxidized low density lipoproteins (oxLDLs), suggesting that these mimics expose the same PC epitopes as those found on oxLDLs. Results from this study suggest that membrane curvature could be an additional factor influencing CRP binding of damaged membranes distinct from the oxidation of PC lipids.
Soft Matter 06/2012; · 4.39 Impact Factor
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ABSTRACT: Here, we demonstrate that aptamers tethered to gold nanoparticles enable direct visualization of protein-oligonucleotide interactions during gel electrophoresis. This technique is used to confirm that an aptamer previously identified as binding to C-reactive protein (CRP) only binds to the monomeric form of CRP. While native, pentameric CRP (pCRP) is used in clinical assays to predict cardiovascular disease (CVD) risk, it is the monomeric isoform that is more strongly associated with pro-inflammatory and pro-atherogenic effects. To visualize this selectivity, the CRP-aptamer was conjugated to streptavidin-coated gold nanoparticles and the mobility of the free oligonucleotide-nanoparticle conjugate (ON-NP) and the protein/ON-NP complex bands were visualized and recorded during electrophoresis using a simple digital camera. At a concentration of 6 μg/mL, monomeric CRP showed a significant decrease in the observed ON-NP mobility, whereas no change in mobility was observed with pCRP up to 18 μg/mL. Advantages of this nanoparticle-based electrophoretic mobility shift assay (NP-EMSA) over the traditional EMSA include real-time detection of protein-oligonucleotide interactions, the avoidance of harmful radioisotopes, and elimination of the need for expensive gel imagers. The availability of both the NP-EMSA technique and an mCRP-specific probe will allow for improved clinical diagnostic to more accurately predict future CVD risk.
Electrophoresis 12/2011; 33(2):348-51. · 3.30 Impact Factor
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ABSTRACT: Native C-reactive protein (CRP) is composed of five identical subunits arranged in a pentameric structure (pCRP). Binding of pCRP to damaged cell membranes produces a second isoform, modified CRP, which has similar antigenicity to isolated monomeric subunits of CRP (mCRP). Emerging evidence indicates that modified CRP plays a role in inflammation and atherosclerosis, however, there are very few techniques that can distinguish the different isoforms of CRP. Here we show that an RNA aptamer binds specifically to mCRP and not to pCRP. Using this aptamer, we describe a simple, fast, and sensitive assay to detect nanomolar concentrations of mCRP using fluorescence anisotropy. In addition, we show that this aptamer can be used to detect mCRP in polyacrylamide gels and bound to a surface using total internal reflection fluorescence microscopy. The biological activity of the mCRP we prepared by heating pCRP with 0.1% sodium dodecyl sulfate was confirmed by observing binding to the complement protein, C1q. This probe provides an important tool for CRP research and has the potential to improve clinical diagnostics that predict risk for cardiovascular disease.
Analytical and Bioanalytical Chemistry 07/2011; 401(4):1309-18. · 3.78 Impact Factor
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ABSTRACT: Lipoprotein particles (LPPs) are biological nanoparticles whose physiological roles are greatly influenced by their sizes. The four major classes of LP are: very low density lipoprotein, intermediate density lipoprotein, low density lipoprotein (LDL) and high density lipoprotein. Since the predominance of small, dense LDLs is associated with increased risk of coronary artery disease (CAD) and diabetes mellitus, LPP profiling can be used to predict metabolic risk factors. Highly tunable LPP mimics can be synthesized using nanoparticles to carefully control for size, lipid composition and surface charge to facilitate the study LPPs in CAD. Here, we engineered LPP mimics using gold nanoparticles between 10-50 nm in diameters. We measured the mobility and zeta potential of these LPP mimics and showed that each mimics have distinct electrokinetic properties and are electrostatically stable.
Proceedings of the ... IEEE Conference on Nanotechnology / co-sponsored by IEEE Robotics and Automation Society (RAS) ... [et al.]. IEEE Conference on Nanotechnology. 01/2011;
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ABSTRACT: Through a careful analysis of the role of formaldehyde in the preparation of silver nanomaterials, a previously unnoticed function of the reagent has been discovered. Formaldehyde reacts with ammonium hydroxide to form a polymer that changes how silver attaches to and coats a substrate. In the case of a gold nanoparticle substrate, this polymer is responsible for creating a nonconcentric core−shell nanoparticle with a near-infrared plasmon resonance at 700 nm. In contrast, when conditions do not favor synthesis of this polymer, concentric nanoparticles are formed that have a plasmon resonance between that of gold and silver at 498 nm. Understanding this second role of formaldehyde allowed us to decrease the amount used 100-fold compared to previous methods, providing a greener synthesis. In addition, it is shown that ascorbic acid can function as a partial substitute for formaldehyde in this synthesis. This strategy may be effective at minimizing or eliminating formaldehyde from the synthesis of other core−shell nanoparticles and nanoshells, facilitating their use in medical applications.
05/2010;
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ABSTRACT: Lipid-coated metal nanoparticles are developed here as a mimic of low-density lipoprotein (LDL) particles and used to study C-reactive protein (CRP) binding to highly curved lipid membranes. A 12 nm shift in the localized surface plasmon resonance (LSPR) was observed when CRP was added to the lipid-coated gold nanoparticles. Transmission electron microscopy (TEM) revealed that CRP induced a structural change to the lipids, resulting in clusters of nanoparticles. This clustering provides a visualization of how CRP could cause the aggregation of LDL particles, which is a key step in atherosclerosis. The cluster formation and resultant LSPR shift requires the presence of both CRP and calcium. Fluorescence anisotropy, using a CRP-specific, fluorophore-labeled aptamer confirmed that CRP was bound to the lipid-coated nanoparticles. An increase in the fluorescence anisotropy (Delta r = +0.261 +/- 0.004) of the aptamer probe occurs in the presence of CRP, PC-coated nanoparticles, and calcium. Subsequent sequestration of calcium by EDTA leads to a decrease in the anisotropy (Delta r = -0.233 +/- 0.011); however, there is no change in the LSPR and no change to the cluster structure observed by TEM. This indicates that CRP binds to the PC membrane on the nanoparticle surface reversibly through a calcium bridging mechanism while changing the underlying membrane structure irreversibly as a result of binding.
The Journal of Physical Chemistry B 04/2010; 114(16):5556-62. · 3.70 Impact Factor
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ABSTRACT: A 1:1 geometrically oriented encounter complex between thieno[2,3-b]pyridine (1) and 4-nitrophenyldia-zoacetate (2) is proposed to account for the dominant formation (ca. 64%) of the 2-isomer in the mixture of 4-nitrophenyl-l isomers obtained previously. A mechanism involving one-electron transfer from 1 to 2 plus fragmentation of 2· into 4-nitrophenyl free radical, N2, and acetate ion is invoked. Formation of other isomers is discussed.It is noted that there is a close correlation between orientational rules plus mechanisms of reaction for numerous free-radical substitutions (SR) with SN reactions of alkyllithiums on furan, thiophene, N-alkylpyrroles, pyridine, and their condensed aromatic molecules, including 1, as substrates. Also isomeric selectivities for SE, SN, and SR substitutions into 1 were shown to be qualitatively consistent with one another. While SE reactions occur largely at position 3 and then at 2, SN and SR reactions occur either at 2 or 6. Selectivity for positions 4 or 5 is small or zero.
Journal of Heterocyclic Chemistry 03/2009; 38(6):1433 - 1439. · 1.22 Impact Factor
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ABSTRACT: Fluorophores from the hemolymph of yellow sac spiders (Cheiracanthium mildei) have been characterized using excitation emission matrix (EEM) fluorescence spectroscopy. This approach provides characterization of fluorophores present in the organism without having to isolate pure samples. Minimal variation occurs between individual samples and each EEM has two distinct peaks, suggesting two fluorophores may be present in the hemolymph. Parallel factor analysis reveals that three fluorophores (with excitation and emission maxima at 270/319, 330/389, and 350/465 nm) best explains the sample to sample variation. By comparing the spectra of the three individual components to fluorophores found in scorpions it is shown that these spiders possess different fluorophores than scorpions. Furthermore, the fluorescence observed is not consistent with beta-carboline or 4-methyl-7-hydroxycoumarin, two compounds previously described in scorpions.
Journal of Photochemistry and Photobiology B Biology 09/2008; 93(3):149-54. · 2.81 Impact Factor
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ABSTRACT: Hybrid bilayers composed of the lipid phosphatidylcholine (PC) and a submonolayer of 1-decanethiol bound to gold nanoparticles are very stable to potassium cyanide.
Chemical Communications 08/2008; · 6.17 Impact Factor
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ABSTRACT: Phosphatidylcholine (PC) is a versatile ligand for synthesizing gold nanoparticles that are soluble in either organic or aqueous media. Here we report a novel route to organic-soluble, PC-stabilized gold nanoparticles that can be re-suspended in water after removal of the organic solvent. Similarly, we show that PC-stabilized gold nanoparticles synthesized in water can be re-suspended in organic solvents after complete removal of water. Without complete removal of the solvent, the nanoparticles retain their original solubility and do not phase transfer. This change in solvent preference from organic to aqueous and vice versa without the use of an additional phase transfer reagent is novel, visually striking, and of utility for synthetic modification of nanoparticles. This approach allows chemical reactions to be performed on nanoparticles in organic solvents followed by conversion of the products to water-soluble materials. A narrow distribution of PC-stabilized gold nanoparticles was obtained after phase transfer to water as characterized by UV-visible (UV-vis) spectroscopy and transmission electron microscopy (TEM), demonstrating that the narrow distribution obtained from the organic synthesis is retained after transfer to water. This method produces water-soluble nanoparticles with a narrower dispersity than is possible with direct aqueous synthesis.
Nanotechnology 03/2008; 19(11):115607. · 3.98 Impact Factor
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ABSTRACT: A porphyrin that contains a single carboxylic acid group was synthesized and coupled to 2'-amino-2'-deoxyuridine. The resultant product contained a free 3' hydroxyl group and a 4,4'-dimethoxytrityl (DMT) protecting group on the 5' hydroxyl of the uridine, making it suitable for use in oligonucleotide synthesis. The 3' H-phosphonate derivative of this molecule was synthesized and used to form a conjugate with a 19 nucleotide sequence of DNA (5'-CCTCCAGTGGAAATCAAGG-3'). This was carried out with the DNA attached at the 3' end to a control pore glass (CPG) substrate, allowing for rapid purification. After removal of the DMT group, an additional three nucleotides were added, leaving the porphyrin as an internal modification. This is the first report of porphyrin attached internally to an oligonucleotide using a hydrogen-bonding nucleoside analog. This allows oligonucleotides to be used as a scaffold for precise positioning of multiple porphyrins within biomimetic arrays.
Bioorganic & medicinal chemistry letters 02/2008; 18(2):850-5. · 2.65 Impact Factor
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ABSTRACT: The evolution of fluorescence is largely unexplored, despite the newfound occurrence of this phenomenon in a variety of organisms. We document that spiders fluoresce under ultraviolet illumination, and find that the expression of this trait varies greatly among taxa in this species-rich group. All spiders we examined possess fluorophores in their haemolymph, but bright fluorescence appears to result when a spider sequesters fluorophores in its setae or cuticle. By sampling widely across spider taxa, we determine that fluorescent expression is labile and has evolved multiple times. Moreover, examination of the excitation and emission properties of extracted fluorophores reveals that spiders possess multiple fluorophores and that these differ among some families, indicating that novel fluorophores have evolved during spider diversification. Because many spiders fluoresce in wavelengths visible to their predators and prey (birds and insects), we propose that natural selection imposed by predator-prey interactions may drive the evolution of fluorescence in spiders.
Biology letters 07/2007; 3(3):265-7. · 3.76 Impact Factor
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ABSTRACT: In this paper we describe a systematic study comparing the properties of self-assembled monolayers (SAMs) formed by in situ deprotection and assembly of S-triphenylmethyl (trityl)- and thiolacetate-protected alkanethiols to those of SAMs formed from the parent alkanethiols. The two in situ deprotections were carried out in trifluoroacetic acid and THF/ammonium hydroxide, respectively. Monolayers of octadecanethiol (ODT) and the peptide-containing alkanethiol 3-mercapto-N-n-pentadecylpropionamide (1ATC15) were assembled on gold using the two in situ methods and characterized by contact angle goniometry, X-ray photoelectron spectroscopy, polarization modulation infrared reflection absorption spectroscopy, and electrochemical characterization methods to assess how the monolayer properties compare to those of monolayers prepared by traditional methods. The results for the in situ deprotection of the trityl-protected molecules demonstrate that this method can afford high-quality monolayers that are nearly indistinguishable from those prepared directly from alkanethiols. The quality of the monolayers prepared using this method is shown to depend on the solubility of the trityl-protected compound in trifluoroacetic acid. The results for the in situ deprotection of acetyl-ODT indicate this method yields low-quality monolayers that contain mixtures of adsorbates bound as thiolates and thiolacetates. In situ trityl deprotection is a useful approach for monolayer formation that greatly simplifies the purification, handling, and assembly of thiol-containing monolayer precursors.
Langmuir 11/2004; 20(21):9144-50. · 4.19 Impact Factor
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ABSTRACT: We have investigated the role of internal functionality in self-assembled monolayers of a family of amide-containing alkanethiol molecules on Au{111} using scanning tunneling microscopy. In addition to van der Waals interactions that are present within n-alkanethiol self-assembled monolayers, hydrogen bonding between adjacent buried amide groups contributes to the stability of the amide-containing molecules on the surface and causes spontaneous phase separation upon coadsorption with an n-alkanethiol. A deposition solution concentration dependence study reveals that this is an observed trend across a range of examined solution compositions. Additionally, hydrogen bonding affects the packing structure of the amide-containing alkanethiol self-assembled monolayers. Although they adopt the same (√3×√3)R30° base lattice as n-alkanethiolate self-assembled monolayers, the amide-containing molecules form superlattice structures that are more linear than n-alkanethiol monolayers due to the hydrogen bonds they form. The internal functionality of monolayers can be used to control their formation and stability.
10/2001;
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ABSTRACT: We report the phase separation of a self-assembled monolayer formed from a binary mixture of adsorbates, n-decanethiol, and an amide-containing alkanethiol of similar length (3-mercapto-N-nonylpropionamide), as studied by scanning tunneling microscopy. While mixtures of n-alkanethiols of similar length (i.e., n-decanethiol and n-dodecanethiol) show no phase separation, the introduction of a hydrogen-bonding functionality buried deep within the film induces the formation of single-component domains on the nanometer scale. Phase separation occurs at all relative compositions studied, and for these molecules maintains the same exposed terminal functionality across the entire film. In nonequimolar concentrations of adsorbates, we observe that the solution component present in greater concentration will dominate the composition of the adsorbed monolayer in super proportion to that in solution, consistent with enthalpic contributions from both the solvent and intermolecular interactions of adsorbates.
02/2001;
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12/2000;
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ABSTRACT: Environmentally benign ("green") chemical techniques are growing in importance in academic and industrial research laboratories. Such chemistry has been slow to appear in teaching laboratories, owing in part to a lack of published material on this subject. Recent developments in green synthesis provide opportunities to introduce this material in teaching laboratories. We present a synthesis of adipic acid that utilizes green reagents (hydrogen peroxide as the oxidant), solvents (water), and methods (phase-transfer catalysis, catalyst recycling). The synthesis works well and provides an excellent forum for emphasizing green chemical concepts while teaching laboratory skills. It demonstrates reuse of a product, synthesis using a nonhazardous solvent, elimination of deleterious by-products, and use of a recyclable catalyst. It can be carried out on either the macroscale or microscale and generates little waste if the catalyst solution is recycled. This experiment fits well in a sophomore organic sequence; it covers the topics of oxidation, phase-transfer catalysis, and the technique of recrystallization, reinforces lecture topics such as alkene synthesis and reactivity, and provides an opportunity to introduce polymer chemistry. Keywords (Domain): Organic Chemistry
11/2000;
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ABSTRACT: A general synthetic approach leading to well-defined, water-soluble gold nanoparticles is described that involves a simple, interfacial ligand exchange reaction between a 1.4 nm phosphine-passivated precursor and an anionic or cationic thiol-containing ligand. We demonstrate the utility of this route by synthesizing water-soluble gold nanoparticles that are stabilized by either an anionic ligand (2-mercaptoethanesulfonate), a cationic ligand (2-(dimethylamino)ethanethiol hydrochloride), or a mixture of both ionic and phosphine ligands. Although the course of the ligand exchange process depends on the nature of the incoming ligand, each of these nanoparticle products retain the small core size and narrow size distribution of the starting particle (1.4 ± 0.4 nm). The stabilities of these nanoparticles to elevated temperature, extremes of pH, and added salt are reported and found to depend on the nature of the exposed headgroups on the ligand shell. Salt-induced aggregation is not observed in any of the cases investigated. Resistance to aggregation is attributed to the protective nature of the ligand shell.
10/2000;
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ABSTRACT: Hybrid self-assembled monolayers (SAMs) containing well-defined strata of different polarity enable insight into how fundamental interactions lead to higher order structure and may provide useful analogies for self-assembled multilayers, new hybrid materials, and functional biological interfaces. We report amide-containing alkanethiol SAMs with internal polar sublayers that are two amide groups thick and nonpolar overlayers comprising either dodecyl or hexadecyl chains. The assemblies have been characterized by X-ray photoelectron spectroscopy (XPS), contact angle goniometry, and external reflective infrared spectroscopy (FTIR-ERS). XPS demonstrates the SAMs are of monolayer thickness, chemisorbed to the gold substrate, and anisotropically oriented. Contact angle data show the methyl surface for n = 16 is highly ordered, but the surface for n = 12 is less well ordered. FTIR-ERS reveals that the alkyl chains for n = 16 are close packed, but that those for n = 12 are disordered. FTIR-ERS also shows that, although the two-amide sublayers are compositionally identical, they are well ordered and assume polyglycine-II-like conformations for n = 16, but they are poorly ordered for n = 12. Comparison of these two SAMs to each other in the context of previously reported one- and three-amide SAMs leads to two conclusions. (1) The threshold n for alkyl chain length ordering in two-amide SAMs is 12 ≤ n ≤ 16. Thus, in SAMs with internal amide sublayers both one and two amide groups thick, the threshold number of methylenes required to form ordered alkyl regions is significantly increased compared to alkanethiol SAMs, demonstrating destructive interference of the amide region with the hydrocarbon ordering process. (2) In two-amide SAMs the formation of a well-ordered amide region depends on the ordering of an overlying hydrocarbon region. This behavior differs with that previously demonstrated for one- and three-amide SAMs, in which the amide groups assume characteristic conformations regardless of hydrocarbon region thickness and order. For two-amide SAMs, the apparent dependence of amide ordering on complementary ordering in the alkyl region provides evidence of an energetic interplay between the two sublayers, manifested as a “reverse ordering” effect. The previously unobserved elastic−elastic character of the buried interface in two-amide SAMs is contrasted with the rigid−elastic interface found in the one-amide SAMs.
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