Scientific publications

Essential for the biological function of the light-driven proton pump, bacteriorhodopsin (BR), and the light sensor, sensory rhodopsin II (SRII), is the coupling of the activated retinal chromophore to the hosting protein moiety. In order to explore the dynamics of this process we have performed ultrafast transient mid-infrared spectroscopy on isotopically labeled BR and SRII samples. These include SRII in D2O buffer, BR in H218O medium, SRII with 15N-labeled protein, and BR with 13C1413C15-labeled retinal chromophore. Via observed shifts of infrared difference bands after photoexcitation and their kinetics we provide evidence for nonchromophore bands in the amide I and the amide II region of BR and SRII. A band around 1550 cm-1 is very likely due to an amide II vibration. In the amide I region, contributions of modes involving exchangeable protons and modes not involving exchangeable protons can be discerned. Observed bands in the amide I region of BR are not due to bending vibrations of protein-bound water molecules. The observed protein bands appear in the amide I region within the system response of ca. 0.3 ps and in the amide II region within 3 ps, and decay partially in both regions on a slower time scale of 9-18 ps. Similar observations have been presented earlier for BR5.12, containing a nonisomerizable chromophore (R. Gross et al. J. Phys. Chem. B 2009, 113, 7851-7860). Thus, the results suggest a common mechanism for ultrafast protein response in the artificial and the native system besides isomerization, which could be induced by initial chromophore polarization.

The marine bacterium strain MC-1 is a member of the alpha subgroup of the proteobacteria that contains the magnetotactic cocci and was the first member of this group to be cultured axenically. The magnetotactic cocci are not closely related to any other known alphaproteobacteria and are only distantly related to other magnetotactic bacteria. The genome of MC-1 contains an extensive (102 kb) magnetosome island that includes numerous genes that are conserved among all known magnetotactic bacteria, as well as some genes that are unique. Interestingly, certain genes that encode proteins considered to be important in magnetosome assembly (mamJ and mamW) are absent from the genome of MC-1. Magnetotactic cocci exhibit polar magneto-aerotaxis, and the MC-1 genome contains a relatively large number of identified chemotaxis genes. Although MC-1 is capable of both autotrophic and heterotrophic growth, it does not appear to be metabolically versatile, with heterotrophic growth confined to the utilization of acetate. Central carbon metabolism is encoded by genes for the citric acid cycle (oxidative and reductive), glycolysis, and gluconeogenesis. The genome also reveals the presence or absence of specific genes involved in the nitrogen, sulfur, iron, and phosphate metabolism of MC-1, allowing us to infer the presence or absence of specific biochemical pathways in strain MC-1. The pathways inferred from the MC-1 genome provide important information regarding central metabolism in this strain that could provide insights useful for the isolation and cultivation of new magnetotactic bacterial strains, in particular strains of other magnetotactic cocci.

We characterize 10-nm outer diameter multi-walled carbon nanotubes (powder and dispersion), which will be used for the preparation of conductive layers on polycarbonate (PC) substrates. The optical, electrical, and mechanical characterization of the spin-coated deposited layers is shown and compared with results obtained for layers deposited in borosilicate glass substrates. In both glass and plastic, the layers have shown transmittance higher than 78% in the visible range and have passed the tape and pencil standard tests for adherence and hardness, respectively. However, the sheet resistance presented by layers deposited on PC is still much superior to that of layers deposited on glass and sintered at higher temperatures. Nevertheless, the results obtained still allow the use of such layers in antistatic applications.

Nanocomposites of TiO2 and multi-walled carbon nanotubes (MWNTs) were produced by the addition of different concentrations of MWNTs to a TiO2 sol matrix. Conductive coatings were prepared by spin coating the nanocomposite sols on glass substrates and sintering the samples at 300 ° C for 15 min. No crystalline phase of TiO2 was formed at this temperature. The sheet resistance of the coatings was decreased from some hundreds of M Ω/sq to just a few k Ω/sq by the addition of MWNTs to the TiO2 matrix. Moreover, sintering of the coatings in a reducing atmosphere has lead to a lower sheet resistance than that presented by coatings sintered in air. The lowest resistivity of 2.0 x 10-1 Ω.cm was obtained by coatings prepared with 5.0 mg/ml MWNTs in the composite sol. The optical, structural and electrical properties of the coatings were correlated and demonstrated.

ITO (tin doped indium oxide) coatings were produced by gravure printing process on PET and PEN foils. The printing paste consists of ITO nanoparticles, which are dispersed in a solvent and mixed with a binder. By modification of the printing paste, the sheet resistance (R/sq) of the ITO coatings after hardening under UV-irradiation at low temperatures (< 130 °C) could be decreased to 1 kΩ/sq. R/sq could be further reduced down to 0.5 kΩ/sq by heat treatment under forming gas atmosphere (N2/H2), the transmission of the ITO coated foils still being more than 80% in the visible range. The application of these ITO films as a bottom electrode in organic photodiodes (OPDs) is shown, and the current density-voltage characteristics of the OPDs are presented.

The fabrication of organic photodiodes on solution cast indium tin oxide (ITO) bottom electrodes on flexible substrates is described. ITO coatings with a sheet resistance of 2 to 3 kΩ/sq are produced by gravure printing process on PEN (polyethylene 2,6-naphthalate) films. The ITO films are directly used as transparent bottom electrodes for organic photodiodes (OPD). Furthermore, first experiments to integrate one of these OPDs in an all-organic opto-chemical sensor are successfully demonstrated. The implementation of OPDs as detectors in applications such as integrated sensors demands a fabrication of these devices on flexible substrates. For these applications the OPDs on printed ITO on PEN are especially suitable.

Steel foil is an attractive candidate for use as a flexible substrate material for Cu(Inx,Ga1-x)Se2 solar cells (CIGS). It is stable at the high temperatures involved during CIGS processing and is also commercially available. Stainless chromium (Cr) steel is more expensive than Cr-free steel sheets, but the latter are not stable against corrosion. We processed CIGS solar cells on both types of substrates. The main problem arising here is the diffusion of detrimental elements from the substrate into the CIGS absorber layer. The diffusion of iron (Fe) and other substrate elements into the CIGS layer was investigated by Secondary Ion Mass Spectrometry (SIMS). The influence of the impurities on the solar cell parameters was determined by current voltage (JV) and external quantum efficiency (EQE) measurements. A direct correlation between the Fe content in the CIGS layer and the solar cell efficiency was found. The diffusion of Fe could be strongly reduced by a diffusion barrier layer. Thus we could process CIGS solar cells with a conversion efficiency of 12.8% even on Cr-free steel substrate.

Inducer-dependent prokaryotic transcriptional repressor proteins that originally evolved to orchestrate the transcriptome with intracellular and extracellular metabolite pools, have become universal tools in synthetic biology, drug discovery, diagnostics and functional genomics. Production of the repressor proteins is often limited due to inhibiting effects on the production host and requires iterative process optimization for each individual repressor. At the example of the Streptomyces pristinaespiralis-derived streptogramin-dependent repressor PIP, the expression of which was shown to inhibit growth of Escherichia coli BL21*, we demonstrate that the addition of the PIP-specific streptogramin antibiotic pristinamycin I neutralizes the growth-inhibiting effect and results in &gt;100-fold increased PIP titers. The yield of PIP was further increased 2.5-fold by the engineering of a new E. coli host suitable for the production of growth-inhibiting proteins encoded by an unfavorable codon usage. PIP produced in the presence of pristinamycin I was purified and was shown to retain the antibiotic-dependent binding to its operator pir as demonstrated by a fluorescence resonance energy transfer (FRET)-based approach. At the example of the macrolide-, tetracycline- and arsenic-dependent repressors MphR(A), TetR and ArsR, we further demonstrate that the production yields can be increased 2- to 3-fold by the addition of the cognate inducer molecules erythromycin, tetracycline and As3+, respectively. Therefore, the addition of inducer molecules specific to the target repressor protein seems to be a general strategy to increase the yield of this interesting protein class. © 2009 Elsevier Inc. All rights reserved.

Adjustable control of therapeutic transgenes in engineered cell implants after transdermal and topical delivery of nontoxic trigger molecules would increase convenience, patient compliance, and elimination of hepatic first-pass effect in future therapies. Pseudomonas putida DOT-T1E has evolved the flavonoid-triggered TtgR operon, which controls expression of a multisubstrate-specific efflux pump (TtgABC) to resist plant-derived defense metabolites in its rhizosphere habitat. Taking advantage of the TtgR operon, we have engineered a hybrid P. putida-mammalian genetic unit responsive to phloretin. This flavonoid is contained in apples, and, as such, or as dietary supplement, regularly consumed by humans. The engineered mammalian phloretin-adjustable control element (PEACE) enabled adjustable and reversible transgene expression in different mammalian cell lines and primary cells. Due to the short half-life of phloretin in culture, PEACE could also be used to program expression of difficult-to-produce protein therapeutics during standard bioreactor operation. When formulated in skin lotions and applied to the skin of mice harboring transgenic cell implants, phloretin was able to fine-tune target genes and adjust heterologous protein levels in the bloodstream of treated mice. PEACE-controlled target gene expression could foster advances in biopharmaceutical manufacturing as well as gene- and cell-based therapies.

Synthetic biology, the science of engineering complex biological systems with novel functions, is increasingly fascinating researchers across disciplines who gather to design functional biological assemblies in a rational and systematic manner. Although initial success stories were based on reprogramming prokaryotic and lower eukaryotic cells, the design of synthetic mammalian gene circuits is becoming increasingly popular because it promises to foster novel therapeutic opportunities in the not-so-distant future. Here, we discuss the latest generation of mammalian synthetic biology devices assembled to form complex synthetic gene networks, such as regulatory cascades, logic evaluators, hysteretic circuits, epigenetic toggle switches, time-keeping components, drug discovery tools, and "cell phone" units. We further highlight how such circuits could be interconnected to achieve higher-order control networks such as synthetic hormone-like communication systems in animals or synthetic ecosystems with dynamic interspecies crosstalk. © 2009 Elsevier Ltd. All rights reserved.