A step-forward in the characterization and potential applications of solid and liquid oxygen transfer vectors

Silicone oil 20 and 200 cSt, a perfluorocarbon (FC40TM), heptamethylnonane, Kraton, Elvax, and Desmopan were evaluated for their ability to enhance oxygen transfer in stirred tank and airlift reactors (STR and ALR, respectively). None of the vectors tested was either toxic or biodegradable and they exhibited a moderate affinity for oxygen (gas/vector partitioning coefficients K_{\textg/\textv} = C_\textg ·C_\textv^{- 1} K_{{{\text{g}}/{\text{v}}}} = C_{\text{g}} \cdot C_{\text{v}}^{- 1} ranging from 3 to 5.1). FC40 was highly volatile, while KratonTM and ElvaxTM exhibited a low thermal stability, which constitutes a serious handicap for their implementation in fermentations. Silicone oil 20 cSt and Desmopan supported the highest oxygen transfer rates under abiotic conditions in both STR and ALR designs, with enhancement factors of up to 90% and 250%, respectively, compared to control tests (deprived of vector). The fact that these vectors showed the highest K _g/v proved that, besides the classical selection criteria, the in situ hydrodynamic behavior (which affects K _L a) must be considered for vector selection. The use of silicone oil 20 cSt and Desmopan in glucose-supplemented Saccharomyces cerevisiae fermentations resulted in a two- and threefold increase in biomass productions, respectively. The better performance of Desmopan in terms of biomass growth enhancement, together with the absence of the operational problems inherent to the use of liquid vectors (such as intensive foaming, high cost, and difficult solvent recovery), make solid vectors a promising and cost-effective alternative in the future developments of two-phase partitioning bioreactors.     

Identification of a novel family of targets of PYK2 related to Drosophila retinal degeneration B (rdgB) protein

The protein tyrosine kinase PYK2 has been implicated in signaling pathways activated by G-protein-coupled receptors, intracellular calcium, and stress signals. Here we describe the molecular cloning and characterization of a novel family of PYK2-binding proteins designated Nirs (PYK2 N-terminal domain-interacting receptors). The three Nir proteins (Nir1, Nir2, and Nir3) bind to the amino-terminal domain of PYK2 via a conserved sequence motif located in the carboxy terminus. The primary structures of Nirs reveal six putative transmembrane domains, a region homologous to phosphatidylinositol (PI) transfer protein, and an acidic domain. The Nir proteins are the human homologues of the Drosophila retinal degeneration B protein (rdgB), a protein implicated in the visual transduction pathway in flies. We demonstrate that Nirs are calcium-binding proteins that exhibit PI transfer activity in vivo. Activation of PYK2 by agents that elevate intracellular calcium or by phorbol ester induce tyrosine phosphorylation of Nirs. Moreover, PYK2 and Nirs exhibit similar expression patterns in several regions of the brain and retina. In addition, PYK2-Nir complexes are detected in lysates prepared from cultured cells or from brain tissues. Finally, the Nir1-encoding gene is located at human chromosome 17p13.1, in proximity to a locus responsible for several human retinal diseases. We propose that the Nir and rdgB proteins represent a new family of evolutionarily conserved PYK2-binding proteins that play a role in the control of calcium and phosphoinositide metabolism downstream of G-protein-coupled receptors.     

Mapping of the interaction between the immunodominant loop of the ectodomain of HIV-1 gp41 and human complement protein C1q.

The human immunodeficiency virus type 1 transmembrane envelope glycoprotein gp41 has been previously shown to activate the C1 complex of human complement through direct interaction with its C1q subunit. The major interaction site has been located within the gp41 immunodominant region (residues 590-620), and a synthetic peptide overlapping residues 601-613 of gp41 (sequence GIWGCSGKLICTT) was shown to inhibit binding of gp41 to C1q in vitro (Thielens, N.M., Bally, I.M., Ebenbichler, C.F., Dierich, M.P. & Arlaud, G.J. (1993) J. Immunol. 151, 6583-6592). The ectodomain of gp41 (s-gp41) was secreted from the methylotrophic yeast Pichia pastoris and purified by immunoaffinity chromatography. Enzymatic deglycosylation of the recombinant s-gp41 was necessary to allow its in vitro interaction with C1q. A solid-phase competition assay was used to monitor the effect of mutant peptides derived from segment 601-613 of gp41 on the binding of deglycosylated s-gp41 to C1q. Whereas mutation of Ser606 had no effect, replacement of Ile602, Trp603, Lys608, Leu609 and Ile610 by Ala abolished the ability of the resulting peptides to inhibit binding of s-gp41 to C1q, suggesting that these residues participate in the interaction between gp41 and C1q. These findings are discussed in the light of a structural model of the immunodominant loop of gp41. It is proposed that the recognition of gp41 by C1q is driven by hydrophobic interactions, and that the sites of gp41 responsible for interaction with gp120 and C1q partly overlap.     

The Content of Alternative Oxidase of Euglena Mitochondria is Increased by Growth in the Presence of Cyanide and is not Cytochrome o.

ABSTRACT A study of the effect of respiratory inhibitors on O₂ uptake of Euglena gracilis mitochondria, isolated from cells grown in the presence of cyanide or with ethanol as carbon source, was undertaken. The contents of cytochrome c oxidase and alternative oxidase were also determined. Inhibition of respiration by antimycin and cyanide was only partial and it was dependent on the oxidizable substrate used. Succinate oxidation was the most sensitive to cyanide whereas lactate oxidation was the most resistant. Cell growth in the presence of cyanide or with ethanol as carbon source brought about an enhanced content of alternative oxidase without a concomitant increase in cytochrome aa₃ content. However, a correlation between cyanide-resistant respiration and alternative oxidase content was not found. Analysis of heme types in mitochondrial membranes revealed the absence of heme O. The data suggest the presence of an inducible alternative oxidase in Euglena mitochondria which has high resistance to cyanide and contains heme B. A close relationship between Euglena alternative oxidase and bacterial quinol oxidases containing B-type heme is proposed.     

Type and contretype signs in monosomy and trisomy 9p. On a case 46,XY, del (9) (pter yields p12:).

A 15-month-old male with a partial monosomy 9p is reported. The comparative analysis with other cases of 9p monosomy demonstrates a typical phenotype which when compared to that of 9p trisomy, permits the delineation of fifteen "type and contretype" signs.     

Dhrs3a regulates retinoic acid biosynthesis through a feedback inhibition mechanism

Retinoic acid (RA) is an important developmental signaling molecule responsible for the patterning of multiple vertebrate tissues. RA is also a potent teratogen, causing multi-organ birth defects in humans. Endogenous RA levels must therefore be tightly controlled in the developing embryo. We used a microarray approach to identify genes that function as negative feedback regulators of retinoic acid signaling. We screened for genes expressed in early somite-stage embryos that respond oppositely to treatment with RA versus RA antagonists and validated them by RNA in situ hybridization. Focusing on genes known to be involved in RA metabolism, we determined that dhrs3a, which encodes a member of the short-chain dehydrogenase/reductase protein family, is both RA dependent and strongly RA inducible. Dhrs3a is known to catalyze the reduction of the RA precursor all-trans retinaldehyde to vitamin A; however, a developmental function has not been demonstrated. Using morpholino knockdown and mRNA over-expression, we demonstrate that Dhrs3a is required to limit RA levels in the embryo, primarily within the central nervous system. Dhrs3a is thus an RA-induced feedback inhibitor of RA biosynthesis. We conclude that retinaldehyde availability is an important level at which RA biosynthesis is regulated in vertebrate embryos.     

Proteome informatics I: bioinformatics tools for processing experimental data

Bioinformatics tools for proteomics, also called proteome informatics tools, span today a large panel of very diverse applications ranging from simple tools to compare protein amino acid compositions to sophisticated software for large-scale protein structure determination. This review considers the available and ready to use tools that can help end-users to interpret, validate and generate biological information from their experimental data. It concentrates on bioinformatics tools for 2-DE analysis, for LC followed by MS analysis, for protein identification by PMF, by peptide fragment fingerprinting and by de novo sequencing and for data quantitation with MS data. It also discloses initiatives that propose to automate the processes of MS analysis and enhance the quality of the obtained results.     

Mouse genomic DNA sequences homologous to sea urchin TU elements are genetically stable polydispersed repeats useful for analysis of multiple RFLPs.

Segments of the murine genome that hybridize to the inverted repeat regions of the transposable TU elements of sea urchins include tandem repeats of a sequence (CTCC) that encodes the recognition site for the restriction enzyme Mnl1, as do the analogous polypurine/polypyrimidine (pPu/pPy) stretches in humans. The Mnl1-sensitive repeats, which exist as a microsatellite sequence 200-300 bp in length, lack the terminal dyad symmetry characteristic of the TU elements and are structurally and functionally distinct from these elements. DNA fragments containing these repeat units that are isolated from different generations of isogenic (or congenic) mice or from different tissues of genetically identical individuals are indistinguishable by RFLP analysis; however, they show restriction fragment length polymorphism in different strains. This polymorphism appears to reflect DNA sequence changes occurring at sites flanking the repeats rather than variability in the number of repeats. Their genetic stability and occurrence in a wide variety of animal species make the Mnl1 repeats useful in studying genetic variation that has occurred over an evolutionary time scale of greater duration than can be examined conveniently by VNTR analysis.