Co-fermentation of xylose and cellobiose by an engineered Saccharomyces cerevisiae

We have integrated and coordinately expressed in Saccharomyces cerevisiae a xylose isomerase and cellobiose phosphorylase from Ruminococcus flavefaciens that enables fermentation of glucose, xylose, and cellobiose under completely anaerobic conditions. The native xylose isomerase was active in cell-free extracts from yeast transformants containing a single integrated copy of the gene. We improved the activity of the enzyme and its affinity for xylose by modifications to the 5′-end of the gene, site-directed mutagenesis, and codon optimization. The improved enzyme, designated RfCO*, demonstrated a 4.8-fold increase in activity compared to the native xylose isomerase, with a K_m for xylose of 66.7?mM and a specific activity of 1.41?μmol/min/mg. In comparison, the native xylose isomerase was found to have a K_m for xylose of 117.1?mM and a specific activity of 0.29?μmol/min/mg. The coordinate over-expression of RfCO* along with cellobiose phosphorylase, cellobiose transporters, the endogenous genes GAL2 and XKS1, and disruption of the native PHO13 and GRE3 genes allowed the fermentation of glucose, xylose, and cellobiose under completely anaerobic conditions. Interestingly, this strain was unable to utilize xylose or cellobiose as a sole carbon source for growth under anaerobic conditions, thus minimizing yield loss to biomass formation and maximizing ethanol yield during their fermentation.     

Correspondenz

Ohne Zusammenfassung     

Effet d'anesthesiques locaux sur le transfert de mannose dans les membranes microsomiques d'Aspergillus niger van Tieghem

The influence of various local anaesthetics has been studied on the mannose transfer enzymic system which is localized in Aspergillus niger van Tieghem microsomal membranes. The n-alcanes and tertiary amines do not seem to react on the first step of the reaction: i.e. the polyprenylphosphate-mannose biosynthesis. However two amines, the dibucaine and to a lesser extent the tetracaine, inhibit the mannose transfer from the polyprenylphosphate-mannose to the endogenous proteins.     

Correspondenz

Ohne Zusammenfassung     

Isaria cretacea van Beyma

Riassunto E' una breve nota descrittiva diIsaria cretacea van Beyma isolata da terreno di bosco ad Eucalipto.Trattasi di un Ifomicete non ancora segnalato presente in Italia e forse non ancora rinvenuto nel terreno.E' specie auxo-eterotrofa e presenta attività antagonista verso altre specie di funghi.

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Summary The present note deals briefly withIsaria cretacea van Beyma, isolated from Eucaliptus soil. The fungus is a Hyphomycete whose presence in Italy was never noticed before and perhaps it is the first time the it is found in soil. ThisIsaria is an auxo-heterotrophic species and shows a certain antagonistic activity against some fungi.

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Resumo O presente trabalho é uma nota breve descritiva deIsaria cretacea van Beyma, isolada de terreno de um bosque de Eucaliptos. Tratase de um Ifomicete cuja presença na Italia não foi ainda assinalada e talvez seja a primeira vez que é isolada de terreno.E' uma especie auxo-heterotrofica e apresenta uma certa atividade antagonista sobre outros fungos.

     

Hieracium pallidifolium n. sp.

Ohne ZusammenfassungAus dem Nachlasse Knaf's mitgetheilt; geschrieben im J. 1863. Die Pflanze ist im Besitze seines Sohnes, eine Abbildung derselben befindet sich in dem soeben erscheinenden 2. Theile von elakovský's Prodromus der Flora von Böhmen.     

Regulation of NADPH Oxidase Activity in Phagocytes: RELATIONSHIP BETWEEN FAD/NADPH BINDING AND OXIDASE COMPLEX ASSEMBLY*

The X⁺-linked chronic granulomatous disease (X⁺-CGD) variants are natural mutants characterized by defective NADPH oxidase activity but with normal Nox2 expression. According to the three-dimensional model of the cytosolic Nox2 domain, most of the X⁺-CGD mutations are located in/or close to the FAD/NADPH binding regions. A structure/function study of this domain was conducted in X⁺-CGD PLB-985 cells exactly mimicking 10 human variants: T341K, C369R, G408E, G408R, P415H, P415L, Δ507QKT509-HIWAinsert, C537R, L546P, and E568K. Diaphorase activity is defective in all these mutants. NADPH oxidase assembly is normal for P415H/P415L and T341K mutants where mutation occurs in the consensus sequences of NADPH- and FAD-binding sites, respectively. This is in accordance with their buried position in the three-dimensional model of the cytosolic Nox2 domain. FAD incorporation is abolished only in the T341K mutant explaining its absence of diaphorase activity. This demonstrates that NADPH oxidase assembly can occur without FAD incorporation. In addition, a defect of NADPH binding is a plausible explanation for the diaphorase activity inhibition in the P415H, P415L, and C537R mutants. In contrast, Cys-369, Gly-408, Leu-546, and Glu-568 are essential for NADPH oxidase complex assembly. However, according to their position in the three-dimensional model of the cytosolic domain of Nox2, only Cys-369 could be in direct contact with cytosolic factors during oxidase assembly. In addition, the defect in oxidase assembly observed in the C369R, G408E, G408R, and E568K mutants correlates with the lack of FAD incorporation. Thus, the NADPH oxidase assembly process and FAD incorporation are closely related events essential for the diaphorase activity of Nox2.     

Guaraná (Paullinia cupana) as a commercial crop in Brazilian Amazonia

Guaraná (Paullinia cupana) is a woody vine or sprawling shrub native to the central Amazon Basin. The seeds are commercially produced on some 6,000 ha in the state of Amazonas near Manaus. The principal article of commerce is an amber-colored, carbonated soft drink. It is also widely used as a high caffeine stimulant and in local medicines. The plant is monoecious and is damaged by a number of diseases, the most severe being anthracnose. Prospects are excellent for greatly expanded international markets.