Origin and evolution of life on terrestrial planets.

The ultimate goal of terrestrial planet-finding missions is not only to discover terrestrial exoplanets inside the habitable zone (HZ) of their host stars but also to address the major question as to whether life may have evolved on a habitable Earth-like exoplanet outside our Solar System. We note that the chemical evolution that finally led to the origin of life on Earth must be studied if we hope to understand the principles of how life might evolve on other terrestrial planets in the Universe. This is not just an anthropocentric point of view: the basic ingredients of terrestrial life, that is, reduced carbon-based molecules and liquid H(2)O, have very specific properties. We discuss the origin of life from the chemical evolution of its precursors to the earliest life-forms and the biological implications of the stellar radiation and energetic particle environments. Likewise, the study of the biological evolution that has generated the various life-forms on Earth provides clues toward the understanding of the interconnectedness of life with its environment.

The effect of the short wavelength ultraviolet radiation. An extension of biological dosimetry to the UV-C range.

Polycrystalline uracil thin layer can be used as biological dosimeter for assessing exposure to UV radiation. The dimerization and reversion efficiency of the ultraviolet radiation in the UV-B and the UV-C range were quantified on polycrystalline uracil thin layers irradiated with quasi-monochromatic radiation using interference filters of 10nm bandwidth. The dimer formation and monomerization (reversion) dose-effect relations were determined by optical spectroscopy. The decrease of the OD value of the uracil thin layer at 288 nm was taken as a measure of the dimer formation, while the increase of the OD of a completely irradiated (until reaching the saturation level) uracil layer was taken as the sign of the monomerization. The two processes in the UV-B and the UV-C range take place simultaneously, the individual characterization of the dimerization efficiency was performed from the initial slope of the dimerization dose-effect function and an action spectrum for dimerization was constructed in the UV-C range too. The reversion efficiency was found to be practically the same with all of the investigated wavelengths: 200 nm, 210 nm, 220 nm, 230 nm, 240 nm The possible biological relevance of the reversion of dimers are discussed.

Annual solar UV exposure and biological effective dose rates on the Martian surface.

The ultraviolet (UV) environment of Mars has been investigated to gain an understanding of the variation of exposure throughout a Martian year, and link this flux to biological effects and possible survival of organisms at the Martian surface. To gain an idea of how the solar UV radiation varies between different regions, including planned landing sites of two future Mars surface missions, we modelled the total solar UV surface flux throughout one Martian year for two different dust scenarios. To understand the degree of solar UV stress on micro-organisms and/or molecules essential for life on the surface of Mars, we also calculated the biologically effective dose (BED) for T7 and Uracil in relevant wavelength regions at the Martian surface as a function of season and latitude, and discuss the biological survival rates in the presence of Martian solar UV radiation. High T7/Uracil BED ratios indicate that even at high latitudes where the UV flux is significantly reduced, the radiation environment is still hostile for life due to the persisting UV-C component of the flux.

Biological UV dosimeters in simulated space conditions.

Polycrystalline uracil thin layers participate in the phage and uracil response (PUR) experiment, assigned to the biological dosimetry of the extraterrestrial solar radiation on the International Space Station (ISS). In ground based experiments (experiment verification tests), the following space parameters were simulated and studied: temperature, vacuum and short wavelength UV (UV-C, down to 200 nm) radiation. The closed uracil samples proved to be vacuum-tight for 7 days. In the tested temperature range (from -20 to +40 degrees C) the uracil samples are stable. The kinetic of dimer formation (dimerization) and reversion (monomerization) of uracil dimers due to short wavelength UV radiation was detected, the monomerization efficiency of the polychromatic deuterium lamp is higher than that of the germicidal lamp. A mathematical model describing the kinetic of monomerization-dimerization was constructed. Under the influence of UV radiation the dimerization-monomerization reactions occur simultaneously, thus the additivity law of the effect of the various wavelengths is not applicable.

Simulation experiments of the effect of space environment on bacteriophage and DNA thin films.

The main goal of PUR experiment (phage and uracil response) is to examine and quantify the effect of specific space conditions on nucleic acid models. To achieve this an improved method was elaborated for the preparation of DNA and bacteriophage thin films. The homogeneity of the films was controlled by UV spectroscopy and microscopy. To provide experimental evidence for the hypothesis that interplanetary transfer of the genetic material is possible, phage T7 and isolated T7 DNA thin films have been exposed to selected space conditions: intense UVC radiation (lambda=254 nm) and high vacuum (10(-4) Pa). The effects of DNA hydration, conformation and packing on UV radiation damage were examined. Characteristic changes in the absorption spectrum, in the electrophoretic pattern of DNA and the decrease of the amount of PCR products have been detected indicating the photodamage of isolated and intraphage DNA.

Stability of nucleic acid under the effect of UV radiation.

Nucleic acids (combined with protein molecules) are essential constituents of the living systems playing an important role in the early evolution of life as well. A specific feature of these molecules has been found and directly confirmed recently: under the influence of short-wavelength UV radiation bipyrimidine photoproducts (cyclobutane dimers and 6-4 bipyrimidines) are induced and the reversion of them can be provoked by the same photons. However, reversion is preferred by the shorter wavelengths. With increasing ratio of the longer wavelength components of the radiation (using artificial UV sources and solar light on the Earth's surface) the impact of the reversible photoproducts in the harmful biological effect decreases and other photoproducts are dominant. Assuming the photoinduced reactions (dimerisation and reversion) are statistical events, during the irradiation the chance for a number of nucleoprotein molecules to survive the radiation damage can be reality. The theoretical and experimental basis of these assumptions will be discussed in the case of bacteriophage T7 nucleoprotein.

S(+)-4-(1-Phenylethylamino)quinazolines as inhibitors of human immunoglobulin E synthesis: potency is dictated by stereochemistry and atomic point charges at N-1.

Since the pathogenesis of allergic diseases is associated with elevated levels of immunoglobulin E (IgE), we developed a high throughput reporter gene assay in a human B-cell line to screen for low molecular weight IgE inhibitory compounds. Monitoring the IL-4 driven IgE-germline promoter activity (IgE-GLP), we discovered 4-(1-phenylethylamino)qinazolines as potent inhibitors of IgE-germline gene expression. Testing of the individual enantiomers (1, 2) revealed that only the S(+) enantiomer 1 was active. A cell viability assay done in the same cell line in parallel discriminated the dose-dependent inhibition from a general antiproliferative effect. The observed correlation of the inhibitory potencies found in the reporter gene assay with those measured by IgE-ELISA in primary human splenocytes provided evidence that the blockade of IgE synthesis is the direct consequence of IgE-germline gene inhibition, thereby validating the reporter gene assay. Parallel synthesis in solution rapidly provided a series of analogues of compound 1 with modifications in the phenethylamine side chain and the quinazoline core for SAR studies. Increasing the lipophilicity of the arylalkylamine moiety yielded S(+)-4-(1-(2-naphthyl)ethylamino)quinazoline (6) as the most potent inhibitor (IC(50) of 14 nM) while the R(-) enantiomer was again found to be inactive. Within the set of S enantiomers, quantum mechanical calculations revealed that the IgE inhibitory activity can be quantitatively described by the charge at N-1 of the heterocyclic core and to a lesser extent by the molar refractivity. These results demonstrate the importance of electron-deficient fused 4-aminopyrimidines and lipophilic side chains for biological activity. The strong preference for the S configuration of the phenethylamine side chain is remarkable insofar as biological activity for fused 4-(1-phenylethylamino)pyrimidines has been published for the R enantiomers only (EGFR tyrosine kinase inhibition).

The conformational origin of the barrier to the formation of neighboring group assistance in glycosylation reactions: a dynamical density functional theory study.

Static and dynamical Density Functional Theory studies of 2,6-di-O-acetyl-3,4-O-isopropylidene-D-galactopyranosyl cation have shown that this cation can exist in two conformers characterized as (2)S(O) and B(2,5), respectively. The (2)S(O) conformer has the O-2 acyl group equatorial with the carbonyl syn to H-2 and is populated by monocyclic oxocarbenium ions. These conformational features are present in the structurally related glycosyl donor ethyl 2,6-di-O-benzoyl-3,4-O-isopropylidene-beta-D-galactothiopyranoside as determined by X-ray diffraction studies. The B(2,5) conformer has O-2 axial and allows the carbonyl to rotate and close the five-membered ring to form a bicyclic dioxolenium ion. Constraints based on natural internal coordinates were implemented to study this conformational transition. In this way the barrier to interconversion has been determined to be 34 kJ mol(-)(1) with a transition state characterized as (O)S(2) and a pathway involving pseudorotation. Thus, for the first time the structures and energetics of the key ions postulated to be involved in neighboring group assisted glycosylation reactions have been determined.

Monitoring of environmental UV radiation by biological dosimeters.

As a consequence of the stratospheric ozone layer depletion biological systems can be damaged due to increased UV-B radiation. The aim of biological dosimetry is to establish a quantitative basis for the risk assessment of the biosphere. DNA is the most important target molecule of biological systems having special sensitivity against short wavelength components of the environmental radiation. Biological dosimeters are usually simple organisms, or components of them, modeling the cellular DNA. Phage T7 and polycrystalline uracil biological dosimeters have been developed and used in our laboratory for monitoring the environmental radiation in different radiation conditions (from the polar to equatorial regions). Comparisons with Robertson-Berger (RB) meter data, as well as with model calculation data weighted by the corresponding spectral sensitivities of the dosimeters are presented. Suggestion is given how to determine the trend of the increase in the biological risk due to ozone depletion.

Influence of phage proteins on formation of specific UV DNA photoproducts in phage T7.

Phage T7 can be used as a biological UV dosimeter. Its reading is proportional to the inactivation rate expressed in HT7 units. To understand the influence of phage proteins on the formation of DNA UV photoproducts, cyclobutane pyrimidine dimers (CPD) and (6-4)photoproducts ((6-4)PD) were determined in T7 DNA exposed to UV radiation under different conditions: intraphage T7 DNA, isolated T7 DNA and heated phage. To investigate the effects of various wavelengths, seven different UV sources have been used. The CPD and (6-4)PD were determined by lesion-specific antibodies in an immunodot-blot assay. Both photoproducts were HT7 dose-dependently produced in all three objects by every irradiation source in the biologically relevant UV dose range (1-10 HT7). The CPD to (6-4)PD ratios increased with the increasing effective wavelength of the irradiation source and were similar in intraphage T7 DNA, isolated DNA and heated phage with all irradiation sources. However, a significant decrease in the yield of both photoproducts was detected in isolated T7 DNA and in heated phage compared to intraphage DNA, the decrease was dependent on the irradiation source. Both photoproducts were affected the same way in isolated T7 DNA and heated phage, respectively. The yield of CPD and (6-4)PD was similar in B, C-like and A conformational states of isolated T7 DNA, indicating that the conformational switch in the DNA is not the decisive factor in photoproduct formation. The most likely explanation for modulation of photoproduct frequency in intraphage T7 DNA is that the presence of bound phage proteins induces an alteration in DNA structure that can result in an increased rate of dimerization and (6-4)PD production of adjacent based in intraphage T7 DNA.

Biological UV dosimeters in the assessment of the biological hazard from environmental radiation.

To determine the impact of environmental UV radiation, biological dosimeters that weight directly the incident UV components of sunlight have been developed, improved and evaluated in the frame of the BIODOS project. Four DNA-based biological dosimeters ((i) phage T7, (ii) uracil thin layer, (iii) spore dosimeter and (iv) DLR-biofilm) have been assessed from the viewpoint of their biological relevance, spectral response and quantification of their biological effectiveness. The biological dosimeters have been validated by comparing their readings with weighted spectroradiometer data, by comparison with other biological doses, as well as with the determined amounts of DNA UV photoproducts. The data presented here demonstrate that the biological dosimeters are potentially reliable field dosimeters for measuring the integrated biologically effective irradiance for DNA damage.

Assessment of the effects of various UV sources on inactivation and photoproduct induction in phage T7 dosimeter.

The correlation between the biologically effective dose (BED) of a phage T7 biological dosimeter and the induction of cyclobutane pyrimidine dimers (CPD) and (6-4) photoproducts ((6-4)PD) in the phage DNA was determined using seven various UV sources. The BED is the inactivation rate of phage T7 expressed in HT7 units. The CPD and (6-4)PD were determined by lesion-specific monoclonal antibodies in an immunodot-blot assay. The various lamps induced these lesions at different rates; the relative induction ratios of CPD to (6-4)PD increased with increasing effective wavelength of irradiation source. The amount of total adducts per phage was compared to the BED of phage T7 dosimeter, representing the average number of UV lesions in phage. For UVC (200-280 nm radiation) and unfiltered TL01 the number of total adducts approximates the reading; however, UV sources having longer effective wavelengths produced fewer CPD and (6-4)PD. A possible explanation is that although the most relevant lesions by UVC are the CPD and (6-4)PD, at longer wavelengths other photoproducts can contribute to the lethal damage of phages. The results emphasize the need to study the biological effects of solar radiation because the lesions responsible for the lethal effect may be different from those produced by various UV sources.

Phage T7 in biological UV dose measurement.

An experimental method complete with theoretical considerations is presented for the measurement of different biological UV doses. The method is based on the high sensitivity of phage T7 activity to UV light. A precisely determined T7 inactivation action spectrum is presented over a wide optical range (240-514 nm). Using the T7 spectral sensitivity in relation to the minimal erythema dose (MED) and the effective spectral irradiance from solar radiation for the MED, an example is given to determine the MED value based on the measurement of T7 inactivation for a given case. The advantages and applicability of the method are discussed.