A.M. Bittner, M. Knez, X.C. Wu, and K. Kern;
M. Sumser, C. Wege, and H. Jeske (Molecular Biology and Plant Virology, Stuttgart University) The fabrication of nanostructures with
well-defined chemical composition and low defect concentration is a prerequisite for the deter-mination of their intrinsic physical proper-ties. However, nanostructures are usually much larger than molecules and they do show defects and thus non-uniformity. Chemically spoken, a macroscopic sample of nanoobjects consists of a distribution of different molecular species.
Directly connected with the size is the problem of uniform chemical composition. The best ex-amples are probably proteins where a change in a single amino acid can make the molecule functional or non-functional. This leads to the idea of using biomolecules either as functional nanostructures or as templates for such.
In order to have a large number of chemical functions, but a well-defined and even stable structure we chose the Tobacco Mosaic Virus (TMV). It exclusively attacks tomato and to-bacco plants and is completely harmless for mammals. The virus is produced and investi-gated in cooperation with the Molecular Biol-ogy and Plant VirolBiol-ogy Department at Stuttgart University. TMV is a stable tube-like complex of a helical RNA strand and 2130 coat proteins (see Fig. 17). The proteins, too, are helically ar-ranged; ca. 16.3 units build up one turn. The
size is 300 nm with 18 nm outer and 4 nm inner diameter. The special shape makes this virus an interesting nanoobject, especially as template for reactions.
Figure 17: Model of ca. 6.9 nm of the 300 nm long Tobacco Mosaic Virus (TMV). Yellow: heli-cal RNA, containing the genome. In the heliheli-cally arranged 49 coat proteins (three turns) each amino acid moiety is colored, reflecting the chemical com-plexity.
On one hand, the molecule is so stable that it tolerates temperatures up to 90ÆC and pH-values from 3.5 to 9 and even several organic solvents such as ethanol or aqueous DMSO. On the other hand, the outer surface provides
hy-droxyl and carboxylate functional groups; in Fig. 17 the orange threonine moieties (hydroxyl groups) on the outer surface are obvious. The inner channel is quite similar, but in addition contains the RNA (yellow) and flexible loops of the protein that comprise threonine, but also the light blue glutamine with its primary amide group. This chemical heterogeneity on the nm scale is – in contrast to uniform inorganic nan-otubes or wires – not usually investigated.
We found that the adsorption behavior of a TMV suspension depends strongly on the chemical properties of the outer surface, i.e., on the carboxylate and on the hydroxyl groups.
This can be exploited by tuning the pH-value of a suspension to fit the substrate chemistry. Even covalent linkages to these groups are possible.
The binding to standard surfaces (gold, graphite and mica, oxidized silicon wafers, glass) was investigated with different protocols such as acid treatment or binding to self-assembled monolayers with ‘sticky’ acid chloride terminal groups.
Figure 18: Non-contact Scanning Force Image of TMV adsorbed on highly oriented pyrolytic graphite. A height profile (cut along the green line) shows 18 nm diameter.
With scanning probe techniques (in air and in aqueous solution) a range of apparent heights from 10 nm to the maximum of 18 nm was found. Non-contact Scanning Force Microscopy turned out to be the optimal method for a direct comparison of apparent heights.
The theoretical diameter / height of 18 nm is
only encountered when surface-virus interac-tions are minimized, e.g., on the unreactive and hydrophobic graphite (see Fig. 18; note that many particles aggregate end-to-end, thus form-ing tubes longer than 300 nm).
Figure 19: Non-contact Scanning Force Image of TMV adsorbed on an oxidized silicon wafer. A height profile (cut along the green line) shows 14 nm height which is less than the diameter of the free virus.
In contrast, lower apparent heights are always connected to hydrophilic interfaces or to co-valent binding as in Fig. 19 on a hydrophilic (oxide-covered) silicon wafer. The lower height is due to stronger interaction, e.g., via hydrogen bonds to the surface, leading to a radial com-pression of the virus. In analogy to biological mechanisms such as dissociation of the protein to liberate the RNA in a plant cell, the flexibil-ity of the rod depends on the chemistry of its surrounding, here of the surface.
In order to arrange the particles in regular ar-rays the adsorption protocols can easily be com-bined with MicroContact Printing. First experi-ments showed that the stability of TMV allows a transfer from a microstructured soft polymer stamp to a surface. A new transfer method was developed which relies on an only partially hy-drophilic stamp that is completely covered by several layers of TMV. In this way large ar-eas (several cm2) can be structured in a fast, highly parallel way. Ideal substrates are hy-drophilic (mica or silicon, see Fig. 19) and can thus strongly bind up to a monolayer of the virus.
Chemical modifications of TMV are not re-stricted to very mild conditions as for most bio-materials. We attained binding and reduction of palladium and gold ions with virus suspen-sions, verified by Transmission Electron Mi-croscopy. The thus produced nanoscale metal clusters can act as catalysts, e.g., for the lo-cal electroless deposition of nickel. In detail, we first adsorbed Pd2 from aqueous PdCl24. An important point was counteracting the hy-drolysis of this complex by working at rather low pH-values and/or high chloride concentra-tions, conditions that may easily be detrimen-tal to other biomolecules. After centrifugation we placed the TMV in a Ni2/ hypophosphite electroless deposition bath of roughly neutral pH, and in some cases heated to 50ÆC. Hy-pophosphite proved to be a reductand that did not affect the TMV structure. The samples were then investigated with Transmission Elec-tron Microscopy. The majority of viruses ap-peared unchanged while about a quarter were filled with up to 6 clusters (black in Fig. 20(a), the single TMV of 300 nm length appears grey).
Figure 20: Transmission Electron Micrographs (200 keV) of TMV on a carbon grid. (a) After treat-ment withPdCl24 and electroless deposition from Ni2+H2PO2; (b) after electroless deposition from Ag+ HCHO. Grey: virus, black: metal clusters.
Surprisingly nearly all clusters were found in-side the virus channel. Electron diffraction proved that these clusters contain nickel. A ten-tative interpretation is that the outermost
sur-face of the coat proteins mainly exhibits alkane and alkanol chains, carboxylate and amides, while RNA and a flexible loop of the coat pro-tein form the channel walls. Here amine groups of various chemical nature can interact with Pd2and bind it stronger than the other chemi-cal groups. In this respect we mention that the channel – albeit narrow – should not prevent the diffusion of species; the hydrophilicity of the RNA and the protein loops should allow for a straightforward wetting of the channel by hy-drophilic liquids.
When the ions are reduced and coalesce to Pd clusters, they can act as activation centers for the electroless deposition of metals like nickel, in fact forming Pd / Ni core / shell clusters. It is exiting that other protocols give different re-sults, e.g., a selective coating of the outer sur-face can be achieved with silver and formalde-hyde as shown in Fig. 20(b). Here the availabil-ity of strong coordination of Ag by carboxy-late on the outer surface may be kinetically fa-vored.
These results point towards simple wet chem-ical syntheses of metal tubes and metal nanowires on biotemplates under full conserva-tion of the biochemical funcconserva-tionality. Incorpo-ration of TMV into existing structures can be achieved by chemical means, i.e., covalent link-ages, offering a ‘bottom-up’ structuring. Alter-natively, MicroContact Printing, a ‘top-down’
structuring method, offers a possibility for ar-ranging (metalized) TMV in defined patterns.
Both should allow contacting the virus cally and thus to gain more insight in its electri-cal properties.
Transport
Transport is one of the basic phenomena in solid state science: transport of electricity, trans-port of heat, transtrans-port of matter. The following contributions are examples of our research on transport of ions in interfaces, on phonon and polariton transport in semiconductors, on electron transport in metals and on charge transport in coupled two-dimensional layers.
Some further highlights of transport studies can be found in other chapters of this report, in particular under the headings of ‘Strongly correlated materials’ and ‘Nanostructures’.