A xylan-degrading strain of<Emphasis Type="Italic"> Sulfolobus solfataricus</Emphasis>: isolation and characterization of the xylanase activity

Two strains (O and X₂) of the hyperthermophilic crenarchaeon Sulfolobus solfataricus strain MT4 were selected and isolated for their ability to grow on xylan. O and X₂, grown on media containing oat spelt xylan and birchwood xylan as the sole nutrient source, respectively, produced the same thermostable xylanase that was demonstrated to be inducible in xylan cultures. In an oat spelt medium, S. solfataricus O underwent interesting morphological changes in the cell envelope, exhibiting mobile appendages not present in the typical coccal shape. The enzyme was prevalently membrane associated and showed a molecular mass of approximately 57.0 kDa. It was also highly thermostable, with a half-life of 47 min at 100°C, and exhibited an optimal temperature and pH of 90°C and 7.0, respectively. Xylo-oligosaccharides were the enzymatic products of xylan hydrolysis, and the smallest degradation product was xylobiose, thus indicating that the enzyme was an endoxylanase. The enzyme was able to bind weakly to crystalline cellulose (Avicel) and more strongly to insoluble xylan in a substrate amount-and temperature-dependent manner.Communicated by G. Antranikian     

Elemental and biochemical composition of<Emphasis Type="Italic"> Nephrops norvegicus</Emphasis> (Linnaeus 1758) larvae from the Mediterranean and Irish Seas

The Norway lobster, Nephrops norvegicus, is a commercially exploited decapod which is widely distributed throughout the north-eastern Atlantic and the Mediterranean Sea. Ovigerous females originating from the Mediterranean and the Irish Seas were held in the laboratory until larvae hatched. Biomass and biochemical composition, as well as digestive gland structure, were examined in newly hatched larvae from these two regions. In addition, previously published data from a North Sea population were included in our comparison. Elemental analyses showed that the absolute quantities of dry mass (DM), carbon (C), nitrogen (N) and hydrogen (H) (collectively referred to as CHN) per individual, and the C:N mass ratios, were significantly lower, while the relative CHN, protein and lipid values (in % of DM) were higher in samples from the Irish Sea compared to larvae originating from either the Mediterranean or the North Sea. As in CHN, the absolute level of protein per individual was higher in larvae from the Mediterranean, while no significant differences were observed in the individual lipid contents. Likewise, the digestive gland structure at hatching did not show any differences between study areas. Intraspecific variability in biomass and chemical composition of newly hatched larvae from different regions may be related to differential patterns of reproduction in regions with different climatic conditions. Lobster larvae hatch in the Mediterranean Sea predominantly in winter when both water temperature and planktonic food availability are at a minimum, while hatching in the Irish Sea occurs under more favourable conditions in spring. Hence, significantly higher wet mass, dry mass and protein values in Mediterranean larvae may represent adaptive traits allowing for early posthatching survival and development under food-limited conditions in an oligotrophic environment.Communicated by H.-D. Franke     

Diversity of the archaeal community in 44 anaerobic digesters as determined by single strand conformation polymorphism analysis and 16S rDNA sequencing

The diversity of Archaea in anaerobic digesters was characterized by strand conformation polymorphism (SSCP) analysis and the sequencing of 16S rDNA genes. The 44 digesters sampled, located in eight different countries, treated effluents from agriculture, the food processing and petro-chemical industries, pulp and paper plant, breweries, slaughterhouses and municipal waste. All the existing processes were represented among the samples (fixed-film, fluidized bed, stirred-tank, UASB, sequential batch reactor, lagoon). Single strand conformation polymorphism analysis targeting the V3 region of 16S rDNA revealed between four to six distinct archaeal peaks per digester. The diversity of dominant Archaea in the 44 digesters was estimated as 23 different 16S rDNA sequences. Cloning of archaeal 16S rRNA genes from 11 distinct total genomic DNA, screening of clones by SSCP and the sequencing of 170 of them made it possible to characterize these SSCP peaks. All the sequences retrieved were members of the Euryarchaeaota subdomain. Furthermore, most of the sequences retrieved were very close to already known and cultivated strains or to environmental clones. The most frequent archaeal sequences were close to Methanosaeta concilii and to a 16S rDNA clone vadinDC06 located in the Methanobacterium clade (84% and 73% of digesters respectively). The other sequences were members of the Methanobacteriales and the Methanomicrobiales families. Only one sequence was far from any sequence of the database and it could be grouped with several sequences of environmental clones. Each digester harboured between two to nine archaeal sequences with only one of them corresponding to a putative acetate-utilizing species. Furthermore, the process in the digesters appeared to play a part in the distribution of archaeal diversity.     

Large-scale exploration of growth inhibition caused by overexpression of genomic fragments in Saccharomyces cerevisiae

We have screened the genome of Saccharomyces cerevisiae for fragments that confer a growth-retardation phenotype when overexpressed in a multicopy plasmid with a tetracycline-regulatable (Tet-off) promoter. We selected 714 such fragments with a mean size of 700 base-pairs out of around 84,000 clones tested. These include 493 in-frame open reading frame fragments corresponding to 454 distinct genes (of which 91 are of unknown function), and 162 out-of-frame, antisense and intergenic genomic fragments, representing the largest collection of toxic inserts published so far in yeast.     

Production of tailor-made fructans in sugar beet by expression of onion fructosyltransferase genes

The consumption of fructans as a low caloric food ingredient or dietary fibre is rapidly increasing due to health benefits. Presently, the most important fructan source is chicory, but these fructans have a simple linear structure and are prone to degradation. Additional sources of high-quality tailor-made fructans would provide novel opportunities for their use as food ingredients. Sugar beet is a highly productive crop that does not normally synthesize fructans. We have introduced specific onion fructosyltransferases into sugar beet. This resulted in an efficient conversion of sucrose into complex, onion-type fructans, without the loss of storage carbohydrate content.     

Nucleotide diversity of the <Emphasis Type="Italic">ZmPox3</Emphasis> maize peroxidase gene: Relationships between a MITE insertion in exon 2 and variation in forage maize digestibility

Background  

Polymorphisms were investigated within the ZmPox3 maize peroxidase gene, possibly involved in lignin biosynthesis because of its colocalization with a cluster of QTL related to lignin content and cell wall digestibility. The purpose of this study was to identify, on the basis of 37 maize lines chosen for their varying degrees of cell wall digestibility and representative of temperate regions germplasm, ZmPox3 haplotypes or individual polymorphisms possibly associated with digestibility.     

Seasonal ovulatory activity exists in tropical Creole female goats and Black Belly ewes subjected to a temperate photoperiod

Background  

Seasonality of ovulatory activity is observed in European sheep and goat breeds, whereas tropical breeds show almost continuous ovulatory activity. It is not known if these tropical breeds are sensitive or not to temperate photoperiod. This study was therefore designed to determine whether tropical Creole goats and Black-Belly ewes are sensitive to temperate photoperiod. Two groups of adult females in each species, either progeny or directly born from imported embryos, were used and maintained in light-proof rooms under simulated temperate (8 to 16 h of light per day) or tropical (11 – 13 h) photoperiods. Ovulatory activity was determined by blood progesterone assays for more than two years. The experiment lasted 33 months in goats and 25 months in ewes.     

Study of protein conformation and orientation in silkworm and spider silk fibers using Raman microspectroscopy

Raman microspectroscopy has been used for the first time to determine quantitatively the orientation of the beta-sheets in silk monofilaments from Bombyx mori and Samia cynthia ricini silkworms, and from the spider Nephila edulis. It is shown that, for systems with uniaxial symmetry such as silk, it is possible to determine the order parameters P2 and P4 of the orientation distribution function from intensity ratios of polarized Raman spectra. The equations allowing the calculation of P2 and P4 using polarized Raman microspectroscopy for a vibration with a cylindrical Raman tensor were first derived and then applied to the amide I band that is mostly due to the C=O stretching vibration of the peptide groups. The shape of the Raman tensor for the amide I vibration of the beta-sheets was determined from an isotropic film of Bombyx mori silk treated with methanol. For both the Bombyx mori and Samia cynthia ricini fibroin fibers, the values of P2 and P4 obtained are equal to -0.36 +/- 0.03 and 0.19 +/- 0.02, respectively, even though the two types of silkworm fibroins strongly differ in their primary sequences. For the Nephila edulis dragline silk, values of P2 and P4 of -0.32 +/- 0.02 and 0.13 +/- 0.02 were obtained, respectively. These results clearly indicate that the carbonyl groups are highly oriented perpendicular to the fiber axis and that the beta-sheets are oriented parallel to the fiber axis, in agreement with previous X-ray and NMR results. The most probable distribution of orientation was also calculated from the values of P2 and P4 using the information entropy theory. For the three types of silk, the beta-sheets are highly oriented parallel to the fiber axis. The orientation distributions of the beta-sheets are nearly Gaussian functions with a width of 32 degrees and 40 degrees for the silkworm fibroins and the spider dragline silk, respectively. In addition to these results, the comparison of the Raman spectra recorded for the different silk samples and the polarization dependence of several bands has allowed to clarify some important band assignments.     

Optimization of the preparation of aqueous suspensions of waxy maize starch nanocrystals using a response surface methodology

Response surface methodology was used to investigate the effect of five selected factors on the selective H(2)SO(4) hydrolysis of waxy maize starch granules. These predictors were temperature, acid concentration, starch concentration, hydrolysis duration, and stirring speed. The goal of this study was to optimize the preparation of aqueous suspensions of starch nanocrystals, i.e., to determine the operative conditions leading to the smallest size of insoluble hydrolyzed residue within the shortest time and with the highest yield. Therefore empirical models were elaborated for the hydrolysis yield and the size of the insoluble residues using a central composite face design involving 31 trials. They allowed us to show that it was possible to obtain starch nanocrystals after only 5 days of H(2)SO(4) hydrolysis with a yield of 15 wt % and having the same shape as those obtained from the classical procedure after 40 days of HCl treatment, with a yield of 0.5 wt %.     

Identification of new intrinsic proteins in Arabidopsis plasma membrane proteome

Identification and characterization of anion channel genes in plants represent a goal for a better understanding of their central role in cell signaling, osmoregulation, nutrition, and metabolism. Though channel activities have been well characterized in plasma membrane by electrophysiology, the corresponding molecular entities are little documented. Indeed, the hydrophobic protein equipment of plant plasma membrane still remains largely unknown, though several proteomic approaches have been reported. To identify new putative transport systems, we developed a new proteomic strategy based on mass spectrometry analyses of a plasma membrane fraction enriched in hydrophobic proteins. We produced from Arabidopsis cell suspensions a highly purified plasma membrane fraction and characterized it in detail by immunological and enzymatic tests. Using complementary methods for the extraction of hydrophobic proteins and mass spectrometry analyses on mono-dimensional gels, about 100 proteins have been identified, 95% of which had never been found in previous proteomic studies. The inventory of the plasma membrane proteome generated by this approach contains numerous plasma membrane integral proteins, one-third displaying at least four transmembrane segments. The plasma membrane localization was confirmed for several proteins, therefore validating such proteomic strategy. An in silico analysis shows a correlation between the putative functions of the identified proteins and the expected roles for plasma membrane in transport, signaling, cellular traffic, and metabolism. This analysis also reveals 10 proteins that display structural properties compatible with transport functions and will constitute interesting targets for further functional studies.