Molecular annotation of ketol-acid reductoisomerases fromStreptomycesreveals a novel amino acid biosynthesis interlock mediated by enzyme promiscuity

The 6-phosphogluconate dehydrogenase superfamily oxidize and reduce a wide range of substrates, making their functional annotation challenging. Ketolacid reductoisomerase (KARI), encoded by the ilvC gene in branched-chain amino acids biosynthesis, is a promiscuous reductase enzyme within this superfamily. Here, we obtain steady-state enzyme kinetic parameters for 10 IlvC homologues from the genera Streptomyces and Corynebacterium, upon eight selected chemically diverse substrates, including some not normally recognized by enzymes of this superfamily. This biochemical data suggested a Streptomyces biosynthetic interlock between proline and the branched-chain amino acids, mediated by enzyme substrate promiscuity, which was confirmed via mutagenesis and complementation analyses of the proC, ilvC1 and ilvC2 genes in Streptomyces coelicolor. Moreover, both ilvC orthologues and paralogues were analysed, such that the relationship between gene duplication and functional diversification could be explored. The KARI paralogues present in S. coelicolor and Streptomyces lividans, despite their conserved high sequence identity (97%), were shown to be more promiscuous, suggesting a recent functional diversification. In contrast, the KARI paralogue from Streptomyces viridifaciens showed selectivity towards the synthesis of valine precursors, explaining its recruitment within the biosynthetic gene cluster of valanimycin. These results allowed us to assess substrate promiscuity indices as a tool to annotate new molecular functions with metabolic implications

Tema 9. Ciclos vitales. Enfoque demográfico

Descripción de la dinámica poblacional. Cómo cuantificar nacimientos, muertes y migraciones

Synthesis of 3-oxo-3-phenyl-2,2,5-trimethyl-1,3-oxaphosphorinanes and their tetrafluoroborate salts

The synthesis and characterization of cis- and trans-3-oxo-3-phenyl-2,2,5-trimethyl-1,3-oxaphosphorinanes (7a and 7b) and their corresponding tetrafluoroborate salts (3a and 3b), heterocyclic organophosphorus compounds not previously reported in the literature, was accomplished. They were fully characterized by H, C and P NMR. It was established the relative configuration of these compounds on the basis of an X-ray diffraction study of oxide 7a

In vitro studies on the effects of fungicides on beneficial fungi of peach twig mycoflora

In vitro studies were carried out to investigate a possible integrated use of chemical and biological means to control the peach twig blight pathogen, Monilinia laxa. Three fungal antagonists of M. laxa (Penicillium purpurogenum, Penicillium frequentans and Epicoccum nigrum) and six fungicides (vinclozolin, iprodione, thiram, captan, benomyl and thiophanate-methyl) were used in the study. Sensitivity of the fungal isolates to the fungicides was determined in vitro by calculating ED50 values. Benomyl and thiophanate-methyl were the most fungitoxic compounds and captan was the least fungitoxic. M. laxa and P. purpurogenum were the most sensitive to all chemicals tested, while E. nigrum and P. frequentans presented bigger differences in their sensitivity to chemicals compared to M. laxa. E. nigrum was consistently less sensitive to benomyl (ED50=2.26 ppm), thiophanate-methyl (ED50=9.61 ppm) and vinclozolin (ED50=3.89 ppm) than the other fungi. P. frequentans was less sensitive to captan, vinclozolin, iprodione, thiophanate-methyl and thiram than M. laxa (8, 7, 5, 4 and 2 times respectively). These results suggest that E. nigrum and P. frequentans could be successfully used in an integrated control programme that combines biological and chemical methods. © 1994 Kluwer Academic Publishers

Involvement of resistance induction by Penicillum oxalicum in the biocontrol of tomato wilt

Penicillum oxalicum, a biocontrol agent for Fusarium oxysporum f.sp. lycopersici, was tested for its ability to induce resistance against tomato wilt. P. oxalicum and F. oxysporum f.sp. lycopersici were placed at separate sites on tomato plants or in soil, avoiding a direct interaction between the fungi. P. oxalicum induced resistance as expressed by a reduction in disease severity, area under disease progress curve and stunting induced by the pathogen. P. oxalicum colonized the tomato rhizosphere during the experiments but it was not detected inside stems, demonstrating that P. oxalicum and Fusarium oxysporum f.sp. lycopersici remained spatially separated. Biological control was observed both in sensitive and 'resistant' cultivars, indicating the role of a general resistance mechanism. In both cultivars P. oxalicum treatment alone did not produce disease symptoms. Therefore P. oxalicum could be a suitable biocontrol agent in cases of cultivar resistance failure. These results suggest that P. oxalicum can trigger defence mechanisms in the plant

Nutritional requirements of antagonists to peach twig blight, Monilinia laxa, in relation to biocontrol

Different carbon and nitrogen sources and accessory substances were tested to determine their effect on the growth and sporulation of the peach twig blight pathogen, Monilinia laxa, and of three of its antagonists (Penicillium frequentans, Penicillium purpurogenum and Epicoccum nigrum), since the success in twig blight biological control by treatments with the fungal antagonists depends on the type of nutrients added to the antagonist formulation. Combinations of sucrose-ammonium tartrate, glucose-(NH4)3PO4-folic acid and lactose-KNO3 were selected from these laboratory experiments because they enhanced the growth and sporulation of P. frequentans, P. purpurogenum and E. nigrum, respectively, but not M. laxa. In glasshouse experiments, twig blight was reduced following the application of mixtures of antagonists with the corresponding enhancing nutrients. © 1993 Kluwer Academic Publishers

A rapid laboratory method for assessing the biological control potential of Penicillium oxalicum against Fusarium wilt of tomato

A rapid, simple and reliable procedure was developed to evaluate biological control of Fusarium wilt of tomato by Penicillium oxalicum. The method consists in growing tomato plants in flasks with nutrient solution in a growth chamber. Plants were previously treated in the seedbed with a conidial suspension (107 conidia mL-1) of P. oxalicum 7 days before transplanting. Fusarium oxysporum f.sp. lycopersici (race 2) was added to the Hoagland solution just before transplanting. Different concentrations and several isolates of F. oxysporum f. sp. lycopersici were tested. Using this method, plants showed typical symptoms of the disease and the effect of the biocontrol agent was clear. Consumption of nutrient solution was reduced in diseased plants, and this reduction was diminished by treatment with P. oxalicum. Consumption of nutrient solution was correlated with other disease-related parameters (AUDPC, weight of aerial parts, stunting) and was an easy and objective parameter to measure

Surface hydrophobicity, viability and efficacy in biological control of Penicillium oxalicum spores produced in aerial and submerged culture

The surface hydrophobicity, viability and biocontrol ability of Penicillium oxalicum spores, produced either in aerial or submerged culture, were characterized. A phase distribution test showed that spores produced in both methods of culture were highly hydrophobic, but those produced in aerial culture were more hydrophobic. Spores stored fresh at either 4 or 25 °C retained a high viability (80%) after 27 weeks of storage, although aerial spores survived better. Freeze-drying severely affected viability, especially of submerged spores. Biocontrol ability against Fusarium oxysporum f. sp. lycopersici was studied in the growth chamber. Aerially- produced spores were more effective than submerged ones. Aerially-produced P. oxalicum spores appeared to have more advantages than those produced by submerged culture, in relation to both viability and efficacy. These results demonstrate that physiological changes occur depending on production conditions which significantly influences quality of the biocontrol agent