Systematic functionalization of molecules for molecular self-assembly

This thesis describes the investigation of systematically varied organic molecules for use in molecular self-assembly processes. All experiments were performed using high-resolution non-contact atomic force microscopy under UHV conditions and at room temperature. Using this technique, three different approaches for influencing intermolecular and molecule-surface interaction on the insulating calcite(10.4) surface were investigated by imaging the structure formation at the molecular scale. I first demonstrated the functionalization of shape-persistent oligo(p-benzamide)s that was engineered by introducing different functional groups and investigating their effect on the structural formation on the sample surface. The molecular core was designed to provide significant electrostatic anchoring towards the surface, while at the same time maintaining the flexibility to fine-tune the resulting structure by adjusting the intermolecular cohesion energy. The success of this strategy is based on a clear separation of the molecule-substrate interaction from the molecule-molecule interaction. My results show that sufficient molecule-surface anchoring can be achieved without restricting the structural flexibility that is needed for the design of complex molecular systems. Three derivatives of terephthalic acid (TPA) were investigated in chapter 7. Here, the focus was on changing the adhesion to the calcite surface by introducing different anchor functionalities to the TPA backbone. For all observed molecules, the strong substrate templating effect results in molecular structures that are strictly oriented along the calcite main crystal directions. This templating is especially pronounced in the case of 2-ATPA where chain formation on the calcite surface is observed in contrast to the formation of molecular layers in the bulk. At the same time, the amino group of 2-ATPA proved an efficient anchor functionality, successfully stabilizing the molecular chains on the sample surface. These findings emphasizes, once again, the importance of balancing and fine-tuning molecule-molecule and molecule-surface interactions in order to achieve stable, yet structurally flexible molecular arrangements on the sample surface. In the last chapter, I showed how the intrinsic property of molecular chirality decisively influences the structure formation in molecular self-assembly. This effect is especially pronounced in the case of the chiral heptahelicene-2-carboxylic acid. Deposition of the enantiopure molecules results in the formation of homochiral islands on the sample surface which is in sharp contrast to the formation of uni-directional double rows upon deposition of the racemate onto the same surface. While it remained uncertain from these previous experiments whether the double rows are composed of hetero- or homochiral molecules, I could clearly answer that question here and demonstrate that the rows are of heterochiral origin. Chirality, thus, proves to be another important parameter to steer the intermolecular interaction on surfaces. Altogether, the results of this thesis demonstrate that, in order to successfully control the structure formation in molecular self-assembly, the correct combination of molecule and surface properties is crucial. This is of special importance when working on substrates that exhibit a strong influence on the structure formation, such as the calcite(10.4) surface. Through the systematic variation of functional groups several important parameters that influence the balance between molecule-surface and molecule-molecule interaction were identified here, and the results of this thesis can, thus, act as a guideline for the rational design of molecules for use in molecular self-assembly.

Local dynamics and bending mechanics of mesostructured materials

Die vorliegende Arbeit behandelt die Anwendung der Rasterkraftmikroskopie auf die Untersuchung mesostrukturierter Materialien. Mesostrukturierte Materialien setzen sich aus einzelnen mesoskopen Bausteinen zusammen. Diese Untereinheiten bestimmen im Wesentlichen ihr charakteristisches Verhalten auf äußere mechanische oder elektrische Reize, weshalb diesen Materialien eine besondere Rolle in der Natur sowie im täglichen Leben zukommt. Ein genaues Verständnis der Selbstorganisation dieser Materialien und der Wechselwirkung der einzelnen Bausteine untereinander ist daher von essentieller Bedeutung zur Entwicklung neuer Synthesestrategien sowie zur Optimierung ihrer Materialeigenschaften. Die Charakterisierung dieser mesostrukturierten Materialien erfolgt üblicherweise mittels makroskopischer Analysemethoden wie der dielektrischen Breitbandspektroskopie, Thermogravimetrie sowie in Biegungsexperimenten. In dieser Arbeit wird gezeigt, wie sich diese Analysemethoden mit der Rasterkraftmikroskopie verbinden lassen, um mesostrukturierte Materialien zu untersuchen. Die Rasterkraftmikroskopie bietet die Möglichkeit, die Oberfläche eines Materials abzubilden und zusätzlich dazu seine quantitativen Eigenschaften, wie die mechanische Biegefestigkeit oder die dielektrische Relaxation, zu bestimmen. Die Übertragung makroskopischer Analyseverfahren auf den Nano- bzw. Mikrometermaßstab mittels der Rasterkraftmikroskopie erlaubt die Charakterisierung von räumlich sehr begrenzten Proben bzw. von Proben, die nur in einer sehr kleinen Menge (<10 mg) vorliegen. Darüberhinaus umfasst das Auflösungsvermögen eines Rasterkraftmikroskops, welche durch die Größe seines Federbalkens (50 µm) sowie seines Spitzenradius (5 nm) definiert ist, genau den Längenskalenbereich, der einzelne Atome mit der makroskopischen Welt verbindet, nämlich die Mesoskala. In dieser Arbeit werden Polymerfilme, kolloidale Nanofasern sowie Biomineralien ausführlicher untersucht.rnIm ersten Projekt werden mittels Rasterkraftmikroskopie dielektrische Spektren von mischbaren Polymerfilmen aufgenommen und mit ihrer lokalen Oberflächenstruktur korreliert. Im zweiten Projekt wird die Rasterkraftmikroskopie eingesetzt, um Biegeexperimente an kolloidalen Nanofasern durchzuführen und so ihre Brucheigenschaften genauer zu untersuchen. Im letzten Projekt findet diese Methode Anwendung bei der Charakterisierung der Biegeeigenschaften von Biomineralien. Des Weiteren erfolgt eine Analyse der organischen Zusammensetzung dieser Biomineralien. Alle diese Projekte demonstrieren die vielseitige Einsetzbarkeit der Rasterkraftmikroskopie zur Charakterisierung mesostrukturierter Materialien. Die Korrelation ihrer mechanischen und dielektrischen Eigenschaften mit ihrer topographischen Beschaffenheit erlaubt ein tieferes Verständnis der mesoskopischen Materialien und ihres Verhaltens auf die Einwirkung äußerer Stimuli.rn

Laserablation von oxidischen Multilagenstrukturen zur Untersuchung von heteroepitaktischen Grenzflächenzuständen

Die Deposition von dünnen, metallischen Schichten auf Silizium-Substraten stellt bereits seit Jahrzehnten die wichtigste Möglichkeit dar, um die wachsenden Anforderungen der Speichertechnologien zu erfüllen. Obwohl Multilagenstrukturen aus oxidischen Schichten eine nahezu unerschöpfliche Vielfalt an neuen Effekten bieten, kommen diese aktuell nur in Nischenanwendungen zum Einsatz. Der Fokus dieser Arbeit liegt auf dem Verständnis von Phänomenen, die nur an Grenzflächensystemen zu beobachten sind. Die Basis der Untersuchungen stellten die Präparation der Multilagenstrukturen durch Laserablation dar. Eine Untersuchung der strukturellen Eigenschaften von multiferroischen BiFeO3 (BFO)-Schichten erlaubte eine Analyse der Wachstumsmodi und der Symmetrie der Einheitszelle von BFO unter heteroepitaktischer Verspannung. Durch Piezokraftmikroskopie konnte die ferroelektrische Domänenstruktur dünner BFO-Schichten analysiert werden. Die Abbildung der magnetischen Domänenstruktur der ferromagnetischen La0,67Sr0,33MnO3 (LSMO)-Schicht und der antiferromagnetischen BFO-Schicht einer Bilagenstruktur durch Photoemissionselektronenmikroskopie erlaubte eine Analyse der Austauschkopplung an der Grenzfläche. Durch elektronische Rekonstruktion entsteht an der LaAlO3 (LAO) /SrTiO3 (STO)-Grenzfläche ein leitfähiger, quasi-zweidimensionaler Zustand. Dessen Transporteigenschaften wurden mit einem Schwerpunkt auf deren Beeinflussung durch ein elektrisches Feld charakterisiert. Diese Ergebnisse führten zur Implementierung einer ferroelektrischen BFO-Schicht zur Manipulation der Leitfähigkeit an der LAO/STO-Grenzfläche. Die Kontrolle des Widerstandes eines mikrostrukturierten Bereichs durch die Polarisation der BFO-Schicht erlaubt die Nutzung der Struktur als Speichertechnologie.

Wet bioadhesion in tree frogs

Viele Tiere wie etwa Geckos oder Laubfrösche können mittels ihrer Haftscheiben an Oberflächen kleben. Diese Haftscheiben ermöglichen es den Tieren, sich während ihrerrnFortbewegung an Oberflächen anzuheften und wieder zu lösen unabhängig von denrnvorherrschenden Umweltbedingungen. Frösche besitzen mikro- und nanostrukturierternsowie charakteristisch geformte Haftscheiben an Finger- und Zehenenden. Ihre besonderernevolutionäre Errungenschaft, sich stark und zugleich reversibel in sowohl trockenen alsrnauch feuchten Umgebungen anzuhaften, hat die Wissenschaft zur Nachahmung und Untersuchungrndieser Strukturen inspiriert. Zum besseren Verständnis der Mechanismen vonrnAnhaftung und Loslösung bei Laubfröschen wurden weiche, elastische und mikrostrukturierternOberflächen hergestellt, indem PDMS (Polydimethylsiloxan) auf einer Siliziummaskernmit Hexagonstruktur aufgetragen und vernetzt wurde. Dadurch wurden Anordnungenrnvon hexagonalen Mikrosäulen mit spezifischen geometrischen Eigenschaften undrnunterschiedlichen Kontaktgeometrien (normale, flache Form, T-Form und konkave Formrnder Säulenenden) erhalten. Um den Einfluss der van-der-Waals, hydrodynamischen,rnKapillar-und Adhäsionskräfte zu verstehen, wurden verschiedene experimentelle Ansätzernverfolgt: Die auf eine einzelne Säule wirkenden Adhäsionskräfte wurden mittelsrnRasterkraftmikroskopie gemessen. Dazu wurden speziell hergestellte kolloidale Sensorenrnverwendet. Diese Experimente wurden sowohl mit als auch ohne Flüssigkeitsfilm auf derrnSäule durchgeführt. Die Ergebnisse zeigten den Beitrag von Kapillarkraft und direktenrnKontaktkräften zur Adhäsionskraft bei Vorliegen eines Flüssigkeitsfilms. Die Adhäsionrnfiel umso größer aus, je weniger Flüssigkeit zwischen Sensor und Säule vorhanden war.rnIm Falle einer trockenen Adhäsion zeigte die Säule mit T-Form die höchste Adhäsion. Darndie Haftscheiben der Laubfrösche weich sind, können sie dynamisch ihre Form ändern,rnwas zu einer Änderung der hydrodynamischen Kraft zwischen Scheibe und Oberflächernführt. Der Einfluss der Oberflächenverformbarkeit auf die hydrodynamische Kraft wurderndaher am Modellsystem einer Kugel untersucht, welche sich einer weichen und ebenenrnOberfläche annähert. Dieses System wurde sowohl theoretisch über die Simulation finiterrnElemente als auch experimentell über die Messung mit kolloidalen Sonden untersucht.rnSowohl experimentelle Ergebnisse als auch die Simulationen ergaben eine Abnahme derrnhydrodynamischen Kraft bei Annäherung des kolloidalen Sensors an eine weiche undrnelastische Oberfläche. Beim Entfernen der Sensors von der Oberfläche verstärkte sichrndie hydrodynamische Anziehungskraft. Die Kraft, die zur Trennung eines Partikels von einer Oberfläche in Flüssigkeit notwendig ist, ist für weiche und elastischen Oberflächenrngrößer als für harte Oberflächen. In Bezug zur Bioadhäsion bei Laubfröschen konnternfestgestellt somit festgestellt werden, dass sich der hydrodynamische Anteil zur feuchtenrnBioadhäsion aufgrund der weichen Oberfläche erhöht. Weiterhin wurde der Einflussrndes Aspektverhältnisses der Säulen auf die Reibungskraft mittels eines kolloidalen Sensorsrnuntersucht. Gestreckte Säulen zeigten dabei eine höhere Reibung im Vergleich zu.rnSäulen mit einem gestreckten Hexagon als Querschnitt.

Stabilizing molecular self-assemblies via on-surface reactions

Für die Realisierung zukünftiger Technologien, wie z.B. molekulare Elektronik, werden Strategien benötigt, um funktionale Strukturen direkt auf Oberflächen zu erzeugen. Für die Bewältigung dieser Aufgabe ist die molekulare Selbstanordnung ein äußerst vielversprechender Bottom-up-Ansatz. Hierbei ist eine der größten Herausforderungen das Zusammenspiel aus intramolekularer Wechselwirkung und der Wechselwirkung zwischen Substrat und Molekülen in ein Gleichgewicht zu bringen. Da jedoch die wirkenden Kräfte der molekularen Selbstanordnung ausschließlich reversibler Natur sind, ist eine langfristige Stabilität fragwürdig. Somit ist die kovalente Verknüpfung der gebildeten Strukturen durch Reaktionen direkt auf der Oberfläche unerlässlich, um die Stabilität der Strukturen weiter zu erhöhen. Hierzu stellt die vorliegende Arbeit eine ausführliche Studie zu molekularer Selbstanordnung und der zielgerichteten Modifikation ebensolcher Strukturen dar. Durch den Einsatz von hochauflösender Rasterkraftmikroskopie im Ultrahochvakuum, welche es erlaubt einzelne Moleküle auf Nichtleitern abzubilden, wurde der maßgebliche Einfluss von Ankerfunktionalitäten auf den Prozess der molekularen Selbstanordnung gezeigt. Des Weiteren konnte die Stabilität der selbst angeordneten Strukturen durch neue Oberflächenreaktionskonzepte entschieden verbessert werden. Der Einfluss von Ankerfunktionen, die elektrostatische Wechselwirkung zwischen Molekül und Substrat vermitteln, auf den Strukturbildungsprozess der molekularen Selbstanordnung wird eingehend durch den Vergleich eines aromatischen Moleküls und seines vierfach chlorierten Derivates gezeigt. Für diese beiden Moleküle wurde ein deutlich unterschiedliches Verhalten der Selbstanordnung beobachtet. Es wird gezeigt, dass die Fähigkeit zur Bildung selbst angeordneter, stabiler Inseln entscheidend durch die Substituenten und die Abmessungen des Moleküls beeinflusst wird. Auch wird in dieser Arbeit die erste photochemische Reaktion organischer Moleküle auf einem Isolator gezeigt. Qualitative und quantitative Ergebnisse liefern ein detailliertes Bild darüber, wie die Abmessungen des Substratgitters die Richtung der Reaktion gezielt beeinflussen. Des Weiteren wird ein allgemeines Konzept zur selektiven Stabilisierung selbstangeordneter Molekülstrukturen durch den kontrollierten Transfer von Elektronen präsentiert. Durch die gezielte Steuerung der Menge an Dotierungsatomen wird die Desorptionstemperatur der molekularen Inseln signifikant erhöht und das Desorptionsverhalten der Inseln entschieden verändert. Diese Arbeit präsentiert somit erfolgreich durchgeführte Strategien um den Prozess der molekularen Selbstanordnung zu steuern, sowie entscheidende Mechanismen um die Stabilisierung und Modifizierung von selbst angeordneten Strukturen zu gewährleisten.

Morphological and optical properties of SiON thin films for photovoltaic applications

In the last years technologies related to photovoltaic energy have rapidly developed and the interest on renewable energy power source substantially increased. In particular, cost reduction and appropriate feed-in tariff contributed to the increase of photovoltaic installation, especially in Germany and Italy. However, for several technologies, the observed experimental efficiency of solar cells is still far from the theoretical maximum efficiency, and thus there is still room for improvement. In this framework the research and development of new materials and new solar devices is mandatory. In this thesis the morphological and optical properties of thin films of nanocrystalline silicon oxynitride (nc-SiON) have been investigated. This material has been studied in view of its application in Si based heterojunction solar cells (HIT). Actually, a-Si:H is used now in these cells as emitter layer. Amorphous SiO_x N_y has already shown excellent properties, such as: electrical conductivity, optical energy gap and transmittance higher than the ones of a-Si:H. Nc-SiO_x N_y has never been investigated up to now, but its properties can surpass the ones of amorphous SiON. The films of nc-SiON have been deposited at the University of Konstanz (Germany). The properties of these films have been studied using of atomic force microscopy and optical spectroscopy methods. This material is highly complex as it is made by different coexisting phases. The main purpose of this thesis is the development of methods for the analyses of morphological and optical properties of nc-SiON and the study of the reliability of those methods to the measurement of the characteristics of these silicon films. The collected data will be used to understand the evolution of the properties of nc-SiON, as a function of the deposition parameters. The results here obtained show that nc-SiON films have better properties with respect to both a-Si:H and a-SiON, i. e. higher optical band-gap and transmittance. In addition, the analysis of the variation of the observed properties as a function of the deposition parameters allows for the optimization of deposition conditions for obtaining optimal efficiency of a HIT cell with SiON layer.

Nano-scale morphological and electrical characterization of nc-SiOxNy thin layers for photovoltaic applications

Il presente lavoro di tesi propone uno studio approfondito di proprietà morfologiche e di trasporto di carica di film sottili di SiOxNy amorfi (a-SiOxNy) e nanocristallini (nc-SiOxNy), che trovano importanti applicazioni in celle fotovoltaiche ad eterogiunzione in silicio, ad alta efficienza. Lo studio è condotto mediante caratterizzazione elettrica e morfologica attraverso tecniche di microscopia a forza atomica (AFM). Sono stati studiati campioni di a-SiOxNy cresciuti con tecnica PECVD (Plasma Enhanced Chemical Vapor Deposition), in cui è stata variata unicamente la distanza tra gli elettrodi durante la deposizione. Sono stati inoltre studiati campioni di nc-SiOxNy, cresciuti con PECVD con una differente percentuale di N2O come gas precursore e un differente tempo di annealing. In entrambi i casi si tratta di un materiale innovativo, le cui proprietà fisiche di base, nonostante le numerose applicazioni, sono ancora poco studiate. L'analisi morfologica, condotta mediante AFM e successiva analisi statistica delle immagini, ha permesso di determinare alcune proprietà morfologiche dei campioni. L’analisi statistica delle immagini è stata validata, dimostrandosi stabile e consistente per lo studio di queste strutture. Lo studio delle proprietà di trasporto è stato condotto mediante acquisizione di mappe di corrente con tecnica conductive-AFM. In questo modo si è ottenuta una mappa di conducibilità locale nanometrica, che permette di comprendere come avviene il trasporto nel materiale. L'analisi di questo materiale mediante tecniche AFM ha permesso di evidenziare che l'annealing produce nei materiali nanocristallini sia un clustering della struttura, sia un significativo aumento della conducibilità locale del materiale. Inoltre la distanza tra gli elettrodi in fase di deposizione ha un leggero effetto sulle dimensioni dei grani. È da notare inoltre che su questi campioni si sono osservate variazioni locali della conducibilità alla nanoscala. L’analisi delle proprietà dei materiali alla nanoscala ha contribuito alla comprensione più approfondita della morfologia e dei meccanismi di trasporto elettronico.

Structure and function of the glucose PTS transporter from Escherichia coli

The glucose transporter IICB of the Escherichia coli phosphotransferase system (PTS) consists of a polytopic membrane domain (IIC) responsible for substrate transport and a hydrophilic C-terminal domain (IIB) responsible for substrate phosphorylation. We have overexpressed and purified a triple mutant of IIC (mut-IIC), which had recently been shown to be suitable for crystallization purposes. Mut-IIC was homodimeric as determined by blue native-PAGE and gel-filtration, and had an eyeglasses-like structure as shown by negative-stain transmission electron microscopy (TEM) and single particle analysis. Glucose binding and transport by mut-IIC, mut-IICB and wildtype-IICB were compared with scintillation proximity and in vivo transport assays. Binding was reduced and transport was impaired by the triple mutation. The scintillation proximity assay allowed determination of substrate binding, affinity and specificity of wildtype-IICB by a direct method. 2D crystallization of mut-IIC yielded highly-ordered tubular crystals and made possible the calculation of a projection structure at 12Å resolution by negative-stain TEM. Immunogold labeling TEM revealed the sidedness of the tubular crystals, and high-resolution atomic force microscopy the surface structure of mut-IIC. This work presents the structure of a glucose PTS transporter at the highest resolution achieved so far and sets the basis for future structural studies.

Immobilization and characterization of recombinant esterase enzyme on chitosan nanoparticles

Immobilization of biologically important molecules on myriad nano-sized materials has attracted great attention. Through this study, thermophilic esterase enzyme was obtained using recombinant DNA technology and purified applying one-step His-Select HF nickel affinity gel. The synthesis of chitosan was achieved from chitin by deacetylation process and degree of deacetylation was calculated as 89% by elemental analysis. Chitosan nanoparticles were prepared based on the ionic gelation of chitosan with tripolyphosphate anions. The physicochemical properties of the chitosan and chitosan nanoparticles were determined by several methods including SEM (Scanning Electron Microscopy), FT-IR (Fourier Transform Infrared Spectroscopy) and DLS (Dynamic Light Scattering). The morphology of chitosan nanoparticles was spherical and the nanospheres’ average diameter was 75.3 nm. The purified recombinant esterase was immobilized efficiently by physical adsorption onto chitosan nanoparticles and effects of various immobilization conditions were investigated in details to develope highly cost-effective esterase as a biocatalyst to be utilized in biotechnological purposes. The optimal conditions of immobilization were determined as follows; 1.0 mg/mL of recombinant esterase was immobilized on 1.5 mg chitosan nanoparticles for 30 min at 60°C, pH 7.0 under 100 rpm stirring speed. Under optimized conditions, immobilized recombinant esterase activity yield was 88.5%. The physicochemical characterization of enzyme immobilized chitosan nanoparticles was analyzed by SEM, FT-IR and AFM (Atomic Force Microscopy).

Fabrication of cone-shaped boron doped diamond and gold nanoelectrodes for AFM-SECM

We demonstrate a reliable microfabrication process for a combined atomic force microscopy (AFM) and scanning electrochemical microscopy (SECM) measurement tool. Integrated cone-shaped sensors with boron doped diamond (BDD) or gold (Au) electrodes were fabricated from commercially available AFM probes. The sensor formation process is based on mature semiconductor processing techniques, including focused ion beam (FIB) machining, and highly selective reactive ion etching (RIE). The fabrication approach preserves the geometry of the original AFM tips resulting in well reproducible nanoscaled sensors. The feasibility and functionality of the fully featured tips are demonstrated by cyclic voltammetry, showing good agreement between the measured and calculated currents of the cone-shaped AFM-SECM electrodes.

Microscopic and Spectroscopic Methods for Probing the Clay Water Interface

Clay minerals have a fundamental importance in many processes in soils and sediments such as the bioavailability of nutrients, water retention, the adsorption of common pollutants, and the formation of an impermeable barrier upon swelling. Many of the properties of clay minerals are due to the unique environment present at the clay mineral/water interface. Traditional techniques such as X-ray diffraction (XRD) and absorption isotherms have provided a wealth of information about this interface but have suffered from limitations. The methods and results presented herein are designed to yield new experimental information about the clay mineral/water interface.A new method of studying the swelling dynamics of clay minerals was developed using in situ atomic force microscopy (AFM). The preliminary results presented here demonstrate that this technique allows one to study individual clay mineral unit layers, explore the natural heterogeneities of samples, and monitor swelling dynamics of clay minerals in real time. Cation exchange experiments were conducted monitoring the swelling change of individual nontronite quasi-crystals as the chemical composition of the surrounding environment was manipulated several times. A proof of concept study has shown that the changes in swelling are from the exchange of interlayer cations and not from the mechanical force of replacing the solution in the fluid cell. A series of attenuated total internal reflection Fourier transform infrared spectroscopy (ATR-FTIR) experiments were performed to gain a better understanding of the organization of water within the interlayer region of two Fe-bearing clay minerals. These experiments made use of the Subtractive Kramers-Kronig (SKK) Transform and the calculation of difference spectra to obtain information about interfacial water hidden within the absorption bands of bulk water. The results indicate that the reduction of structural iron disrupts the organization of water around a strongly hydrated cation such as sodium as the cation transitions from an outer-sphere complex with the mineral surface to an inner-sphere complex. In the case of a less strongly hydrated cation such as potassium, reduction of structural iron actually increases the ordering of water molecules at the mineral surface. These effects were only noticed with the reduction of iron in the tetrahedral sheet close to the basal surface where the increased charge density is localized closer to the cations in the interlayer.

Investigating The Fate Of Positively Charged Nanoparticles In A Soil Environment

The recent increase in the amount of nanoparticles incorporated into commercial products is accompanied by a rising concern of the fate of these nanoparticles. Once released into the environment, it is inevitable that the nanoparticles will come into contact with the soil, introducing them to various routes of environmental contamination. One route that was explored in this research was the interaction between nanoparticles and clay minerals. In order to better define the interactions between clay minerals and positively charged nanoparticles, in situ atomic force microscopy (AFM) was utilized. In situ AFM experiments allowed interactions between clay minerals and positively charged nanoparticles to be observed in real time. The preliminary results demonstrated that in situ AFM was a reliable technique for studying the interactions between clay minerals and positively charged nanoparticles and showed that the nanoparticles affected the swelling (height) of the clay quasi-crystals upon exposure. The preliminary AFM data were complemented by batch study experiments which measured the absorbance of the nanoparticle filtrate after introduction to clay minerals in an effort to better determine the mobility of the positively charged nanoparticles in an environment with significant clay contribution. The results of the batch study indicated that the interactions between clay minerals and positively charged nanoparticles were size dependent and that the interactions of the different size nanoparticles with the clay may be occurring to different degrees. The degree to which the different size nanoparticles were interacting with the clay was further probed using FTIR (Fourier transform infrared) spectroscopy experiments. The results of these experiments showed that interactions between clay minerals and positively charged nanoparticles were size dependent as indicated by a change in the FTIR spectra of the nanoparticles upon introduction to clay.

Reconstitution of mitochondrial ATP synthase into lipid bilayers for structural analysis

Mitochondrial F(1)F(o)-ATP synthase is a molecular motor that couples the energy generated by oxidative metabolism to the synthesis of ATP. Direct visualization of the rotary action of the bacterial ATP synthase has been well characterized. However, direct observation of rotation of the mitochondrial enzyme has not been reported yet. Here, we describe two methods to reconstitute mitochondrial F(1)F(o)-ATP synthase into lipid bilayers suitable for structure analysis by electron and atomic force microscopy (AFM). Proteoliposomes densely packed with bovine heart mitochondria F(1)F(o)-ATP synthase were obtained upon detergent removal from ternary mixtures (lipid, detergent and protein). Two-dimensional crystals of recombinant hexahistidine-tagged yeast F(1)F(o)-ATP synthase were grown using the supported monolayer technique. Because the hexahistidine-tag is located at the F(1) catalytic subcomplex, ATP synthases were oriented unidirectionally in such two-dimensional crystals, exposing F(1) to the lipid monolayer and the F(o) membrane region to the bulk solution. This configuration opens a new avenue for the determination of the c-ring stoichiometry of unknown hexahistidine-tagged ATP synthases and the organization of the membrane intrinsic subunits within F(o) by electron microscopy and AFM.

The supramolecular assemblies of voltage-dependent anion channels in the native membrane

Voltage-dependent anion channels (VDACs) are major constituents of the outer mitochondrial membrane (OMM). These primary transporters of nucleotides, ions and metabolites mediate a substantial portion of the OMM molecular traffic. To study the native supramolecular organization of the VDAC, we have isolated, characterized and imaged OMMs from potato tubers. SDS-PAGE and mass spectrometry of OMMs revealed the presence of the VDAC isoforms POM34 and POM36, as well as the translocase of the OMM complex. Tubular two-dimensional crystals of the VDAC spontaneously formed after incubation of OMMs for two to three months at 4 degrees C. Transmission electron microscopy revealed an oblique lattice and unit cells housing six circular depressions arranged in a hexagon. Atomic force microscopy of freshly isolated OMMs demonstrated (i) the existence of monomers to tetramers, hexamers and higher oligomers of the VDAC and (ii) its spatial arrangement within the oligomers in the native membrane. We discuss the importance of the observed oligomerization for modulation of the VDAC function, for the binding of hexokinase and creatine kinase to the OMM and for mitochondria-mediated apoptosis.

Osteoblast responses to different oxide coatings produced by the sol-gel process on titanium substrates

In order to improve the osseointegration of endosseous implants made from titanium, the structure and composition of the surface were modified. Mirror-polished commercially pure (cp) titanium substrates were coated by the sol-gel process with different oxides: TiO(2), SiO(2), Nb(2)O(5) and SiO(2)-TiO(2). The coatings were physically and biologically characterized. Infrared spectroscopy confirmed the absence of organic residues. Ellipsometry determined the thickness of layers to be approximately 100nm. High resolution scanning electron microscopy (SEM) and atomice force microscopy revealed a nanoporous structure in the TiO(2) and Nb(2)O(5) layers, whereas the SiO(2) and SiO(2)-TiO(2) layers appeared almost smooth. The R(a) values, as determined by white-light interferometry, ranged from 20 to 50nm. The surface energy determined by the sessile-drop contact angle method revealed the highest polar component for SiO(2) (30.7mJm(-2)) and the lowest for cp-Ti and 316L stainless steel (6.7mJm(-2)). Cytocompatibility of the oxide layers was investigated with MC3T3-E1 osteoblasts in vitro (proliferation, vitality, morphology and cytochemical/immunolabelling of actin and vinculin). Higher cell proliferation rates were found in SiO(2)-TiO(2) and TiO(2), and lower in Nb(2)O(5) and SiO(2); whereas the vitality rates increased for cp-Ti and Nb(2)O(5). Cytochemical assays showed that all substrates induced a normal cytoskeleton and well-developed focal adhesion contacts. SEM revealed good cell attachment for all coating layers. In conclusion, the sol-gel-derived oxide layers were thin, pure and nanostructured; consequent different osteoblast responses to those coatings are explained by the mutual action and coadjustment of different interrelated surface parameters.