Bernard Barbier

Survival of microorganisms in space protected by meteorite material: Results of the experiment ‘EXOBIOLOGIE’ of the PERSEUS mission

During the early evolution of life on Earth, before the formation of a protective ozone layer in the atmosphere, high intensities of solar UV radiation of short wavelengths could reach the surface of the Earth. Today the full spectrum of solar UV radiation is only experienced in space, where other important space parameters influence survival and genetic stability additionally, like vacuum, cosmic radiation, temperature extremes, microgravity. To reach a better understanding of the processes leading to the origin, evolution and distribution of life we have performed space experiments with microorganisms. The ability of resistant life forms like bacterial spores to survive high doses of extraterrestrial solar UV alone or in combination with other space parameters, e.g. vacuum, was investigated. Extraterrestrial solar UV was found to have a thousand times higher biological effectiveness than UV radiation filtered by stratospheric ozone concentrations found today on Earth. The protective effects of anorganic substances like artificial or real meteorites were determined on the MIR station. In the experiment EXOBIOLOGIE of the French PERSEUS mission (1999) it was found that very thin layers of anorganic material did not protect spores against the deleterious effects of energy-rich UV radiation in space to the expected amount, but that layers of UV radiation inactivated spores serve as a UV-shield by themselves, so that a hypothetical interplanetary transfer of life by the transport of microorganisms inside rocks through the solar system cannot be excluded, but requires the shielding of a substantial mass of anorganic substances.

THE PERSEUS EXOBIOLOGY MISSION ON MIR: BEHAVIOUR OF AMINO ACIDS AND PEPTIDES IN EARTH ORBIT

Leucine, α-methyl leucine and two peptides were exposed tospace conditions on board the MIR station during the Perseus-Exobiology mission. This long duration space mission was aimed at testing the delivery of prebiotic building blocks. Duringthis mission, two amino acids (leucine and α-methyl leucine) and two peptides (leucine-diketopiperazine and trileucine thioethylester) were exposed in Earth orbit for threemonths. Basalt, clay and meteorite powder were also mixed with the samples in order to simulate the effects of potential meteorite protection. Analysis of the material after the flight did not reveal any racemization or polymerisation but did provideinformation regarding photochemical pathways for the degradationof leucine and of the tripeptide. Amino acids appeared to be moresensitive to UV radiation than peptides, the cyclic dipeptide being found to be as particularly resistant. Meteorite powder which exhibits the highest absorption in Vacuum UltraViolet (VUV)afforded the best protection to the organic molecules whereasmontmorillonite clay, almost transparent in VUV, was the leastefficient. By varying the thickness of the meteorite, we found that the threshold for efficient protection against radiation was about 5 μm. The possible exogenous origin of biologicalbuilding blocks is discussed with respect to the stability to themolecules and the nature of the associated minerals.

Photochemical processing of amino acids in Earth orbit

Abstract Space technology in Earth orbit has been used to investigate whether amino acids and peptides required for the emergence of life can be safely transported to Earth vicinity when they are associated with minerals. In the BIOPAN-1 flight experiment, l -amino acids and one dipeptide were exposed to space conditions, free and associated with clays. Six amino acids found in the Murchison meteorite (Gly, Ala, Leu, Val, Asp, Glu) were tested with respect to chemical degradation and racemization. In addition, photosensitive l -tyrosine was used to check possible oligotyrosine formation. The dipeptide l -alanyl- l -alanine was chosen to test the stability of the peptide bond. No detectable traces of d -amino acids could be found after the flight in any of the samples. Aspartic acid and glutamic acid exposed as free samples have been partially decomposed during exposure to solar UV. Decomposition was prevented when the amino acids were embedded in montmorillonite or kaolinite. The other amino acids were unaffected by the flight. Tyrosine did not condense and the dipeptide remained stable.

Chemical activity of simple basic peptides

Alternating all-L poly(leucyl-lysyl) increases markedly the rate of hydrolysis of oligoribonucleotides. Pure D poly (leucyl-lysyl) is as active as the all-L polymer. The homochiral polypeptides adopt aβ-sheet structure when complexed to the oligonucleotides. Alternating poly(D,L-Leu-D,L-Lys) made of racemic amino acids is much less efficient and is unable to adopt aβ-sheet structure. A set of alternating poly (leucyl-lysyl) ranging from the racemic to the homochiral all-L polymer has been checked. Their conformations can be described as a mixture of random coil andβ-sheet conformations, the amount ofβ-sheet increasing with the optical purity of the polymer. The hydrolytic activity follows the proportion ofβ-sheets, suggesting that the chemical activity is related to the geometry of the chain. Short peptides were prepared in order to evaluate the critical chain length required for the hydrolytic activity. A decapeptide is long enough to present 90% of the activity of the corresponding polypeptide.

Exposure of amino acids and derivatives in the Earth orbit

Abstract Amino acids and amino acid derivatives were exposed to space conditions in Earth orbit as part of the ESA BIOPAN-2 mission to test the possible delivery of extraterrestrial biological building blocks to the primitive Earth. During the Biopan-2 mission, four proteinaceous amino acids (glycine, aspartic acid, glutamic acid and tyrosine), some amino acid esters and two peptides were exposed in Earth orbit for 10 days. Samples were exposed to vacuum and to solar radiation down to 120 nm both alone or associated with montmorillonite as dry films deposited on MgF2 windows. The compounds recovered after the flight were analysed in order to assess chemical degradation, racemization and polymerization. The results confirmed the absence of racemization of the exposed molecules and the high sensitivity of acidic amino acids towards UV radiation already observed in the Biopan-1 exposure mission. Reducing the thickness of the films revealed unexpected sensitivities of exposed amino acids and peptides. A slight protecting effect was observed when the samples were embedded in 5 μm thick montmorillonite films. Several amino acid esters were also exposed to study their possible polymerization in space. Their stability and reactivity in space conditions were compared. Significant degradation was observed for exposed unprotected samples implying that some kind of protection is needed to ensure any amino acid survival in space. Montmorillonite provided some protection but is not an ideal shielding material.

T helper cell epitopes of the human immunodeficiency virus (HIV-1) nef protein in rats and chimpanzees

T helper cell antigenic and immunogenic determinants of the nef protein were investigated in the rat and chimpanzee models using recombinant nef protein and five synthetic peptides selected according to their amphipathic and alpha-helicity properties. The nef protein was shown to be immunogenic with both Freunds or aluminium hydroxide adjuvants. After immunization with the nef protein the 45-69 peptide was the most antigenic in rat and monkey models. In contrast, the 98-112 peptide, that required a carrier protein to induce in vitro rat T cell recall proliferation, was able to restimulate monkey T cells in the absence of a carrier. The amino acid sequence carrying the antigenic activity of the 45-69 peptide was further investigated by synthesizing short peptides overlapping this region. The antigenic sequence was precisely located in the middle of the peptide (region 50-59). This sequence was antigenic only when N alpha-acetylated. Circular dichroism analysis of the 45-69 peptide and the in vitro activity of the N-terminus group indicate in this case the involvement of the alpha-helical propensity for antigen presentation. However, the shorter sequence 50-64, able to induce a T cell reactivity, was determined as a beta-pleated sheet structure in aqueous solution. The 45-69 peptide was not only antigenic but also immunogenic and behaved in vivo as a functional T helper cell epitope. Indeed, the priming with the peptide or the transfer of peptide specific T cells to a naive recipient, followed by immunization with the nef protein, enhanced the subsequent antibody response to the nef protein. Together, these data indicate that the 45-69 peptide appears as a candidate for the in vivo elicitation of T cell immunity to the HIV-1 nef regulatory protein.

Zinc-Induced Conformational Transitions of Acidic Peptides: Characterization by Circular Dichroism and Electrospray Mass Spectrometry

The addition of Zn2+to (Asp-Leu)n and (Leu-Asp-Asp-Leu)n-Asp, which adopt a random coil conformation in pure water, induces a β-sheet and an α-helix structure, respectively. These conformational transitions are dependent on the chain length. The self-association of α-helices as a function of the temperature was studied by CD spectroscopy. The number of Zn2+ ions bound by the peptides was observed to be less than the theoretically expected value.

Circular Polarization of Light by Planet Mercury and Enantiomorphism of its Surface Minerals

Different mechanisms for the generation of circular polarization by the surface of planets and satellites are described. The observed values for Venus, the Moon, Mars, and Jupiter obtained by photo-polarimetric measurements with Earth based telescopes, showed accordance with theory. However, for planet Mercury asymmetric parameters in the circular polarization were measured that do not fit with calculations. For BepiColombo, the ESA cornerstone mission 5 to Mercury, we propose to investigate this phenomenon using a concept which includes two instruments. The first instrument is a high-resolution optical polarimeter, capable to determine and map the circular polarization by remote scanning of Mercurys surface from the Mercury Planetary Orbiter MPO. The second instrument is an in situ sensor for the detection of the enantiomorphism of surface crystals and minerals, proposed to be included in the Mercury Lander MSE.

Improved detection of human antibodies to a Plasmodium antigen using a peptide modified with Aib residues.

A 17‐mer sequence was selected as a model to study the influence of modifications of terminal ends both on the conformation of a peptide and on its antigenicity towards naturally developing antibodies. This sequence corresponded to a tandemly repeated motif, found in a long repetitive region, with high helical propensity, of a Plasmodium falciparum liver‐stage antigen (LSA‐1), immunogenic in man. Our model peptide was synthesized with ionizable or non‐ionizable ends, or modified in both extremities by introduction of the helix‐promoting residue α‐aminoisobutyric acid (Aib). Helical contribution, absent in the 17 amino‐acid sequence possessing ionizable ends, was detectable when non‐ionizable ends were introduced, and dramatically increased in the Aib‐modified analogue. The presence of ionizable ends totally abolished reactivity towards human sera, otherwise detectable with the peptide possessing non‐ionizable ends. While modification by Aib residues was neither detrimental nor beneficial to antigenicity in solution, it clearly resulted in an improved sensitivity of the specific antibody detection when used as solid‐phase antigen in ELISA.

Early peptidic enzymes

Oligopeptides supposed to be essential to primitive living cells could not be obtained by a prebiotic organic chemistry working mainly at random. Selection pathways were required. Experimental evidence is given for selective condensation of amino-acids in water as well as for selective resistance to degradation. Polycationic polypeptides containing lysyl (or arginyl) and hydrophobic residues strongly accelerate the hydrolysis of oligoribonucleotides. A ionic complex is first formed and the polypeptides are particularly active when they adopt a stable conformation, β-sheet or α-helix, in the complex. Well-defined short peptides were synthesized in order to determine the critical chain-length required for chemical activity. In a contemporary cell, proteins represent about 40 % of the dry weight. They fulfil a structural role and they are particularly helpful as chemical catalysts (enzymes). They can be represented as long chains made of twenty different building blocks, the amino-acids NH2CHRCOOH, which differ by the side-chain R. Proteins are remarkable in the sense that they use amino-acids having only one carbon atom between the NH2 and COOH functions. The central carbon atom has always the same spatial asymmetry (chirality) and always bears a hydrogen atom. When the side-chain R is a hydrocarbon, it is branched. When R contains a chemical function, the side functions do not participate to the peptide bond construction. The protein chain results from the condensation of amino-acids, i.e. water molecules are removed between molecules in a medium which is mainly aqueous (the cell contains 75 % of water). The protein chains adopt rigid asymmetric conformations (α-helices, β-sheet structures) which are essential for the protein functions. Proteins, even the smallest ones, are too sophisticated entities to be considered as the products of an organic chemistry working at random, without any chemical selection. The chemist has therefore to understand, with simple models, how primitive proteins were selected and how they began to exhibit chemical activity.