Laura Levin

Layer-by-Layer Self-Assembled Osmium Polymer-Mediated Laccase Oxygen Cathodes for Biofuel Cells: The Role of Hydrogen Peroxide

High potential purified Trametes trogii laccase has been studied as a biocatalyst for oxygen cathodes composed of layer-by-layer self-assembled thin films by sequential immersion of mercaptopropane sulfonate-modified Au electrode surfaces in solutions containing laccase and osmium-complex bound to poly(allylamine), (PAH-Os). The polycation backbone carries the Os redox relay, and the polyanion is the enzyme adsorbed from a solution of a suitable pH so that the protein carries a net negative charge. Enzyme thin films were characterized by quartz crystal microbalance, ellipsometry, cyclic voltammetry, and oxygen reduction electrocatalysis under variable oxygen partial pressures with a rotating disk electrode. New kinetic evidence relevant to biofuel cells is presented on the detection of traces of H(2)O(2), intermediate in the O(2) reduction, with scanning electrochemical microscopy (SECM). Furthermore the inhibitory effect of peroxide on the biocatalytic current resulted in abnormal current dependence on the O(2) partial pressure and peak shape with hysteresis in the polarization curves under stagnant conditions, which is offset upon stirring with the RDE. The new kinetic evidence reported in the present work is very relevant for the operation of biofuel cells under stagnant conditions of O(2) mass transport.

Pectinolytic enzyme production by Colletotrichum truncatum, causal agent of soybean anthracnose

BACKGROUND Colletotrichum truncatum is the most common pathogenic fungus associated with soybean anthracnose, a prevalent disease in Argentina. Pectinolytic enzymes are involved in the pathogenicity of a wide range of plant pathogenic fungi. OBJECTIVES To explore pectinolytic enzyme production in Argentinian Colletotrichum strains isolated from diseased soybean plants from different geographic locations, as a preliminary step to establish the biological role of the pectinolytic enzymes in the Colletotrichum spp.-soybean system, yet unknown. METHODS Ten strains were screened for in vitro pectinolytic enzyme production on a defined medium based on pectin as carbon source. RESULTS All isolates were able to grow in this medium and polymethylgalacturonase (PMG), polygalacturonase (PG) and pectin lyase (PL) activities were detected. On the whole, the peak of polygalacturonases activities preceded the day of maximum growth, while PL activity reached its highest level afterwards. Strain BAFC 3097 (from Santa Fe province) yielded high titles of the three enzymes (1.08U/ml PG, 1.05U/ml PMG, 156U/ml PL), after a short incubation period (7-10 days). Low synthesis of polygalacturonases in cultures containing glucose as unique carbon source suggests that these enzymes are constitutive in contrast with PL, which was not detected. CONCLUSIONS The disparity observed in enzyme production among strains cannot be related to fungal growth, since no major differences in mycelial yield were found; it was not connected with their geographic origin, but might be associated with differences in virulence among strains not yet evaluated.

Genome Sequence of the Melanin-Producing Extremophile Aeromonas salmonicida subsp. pectinolytica Strain 34melT

ABSTRACT The genome of Aeromonas salmonicida subsp. pectinolytica strain 34melT, isolated from a heavily polluted river, contains several genomic islands and putative virulence genes. The identification of genes involved in resistance to different kinds of stress sheds light on the mechanisms used by this strain to thrive in an extreme environment.

Living in an Extremely Polluted Environment: Clues from the Genome of Melanin-Producing Aeromonas salmonicida subsp. pectinolytica 34melT

ABSTRACT Aeromonas salmonicida subsp. pectinolytica 34melT can be considered an extremophile due to the characteristics of the heavily polluted river from which it was isolated. While four subspecies of A. salmonicida are known fish pathogens, 34melT belongs to the only subspecies isolated solely from the environment. Genome analysis revealed a high metabolic versatility, the capability to cope with diverse stress agents, and the lack of several virulence factors found in pathogenic Aeromonas. The most relevant phenotypic characteristics of 34melT are pectin degradation, a distinctive trait of A. salmonicida subsp. pectinolytica, and melanin production. Genes coding for three pectate lyases were detected in a cluster, unique to this microorganism, that contains all genes needed for pectin degradation. Melanin synthesis in 34melT is hypothesized to occur through the homogentisate pathway, as no tyrosinases or laccases were detected and the homogentisate 1,2-dioxygenase gene is inactivated by a transposon insertion, leading to the accumulation of the melanin precursor homogentisate. Comparative genome analysis of other melanogenic Aeromonas strains revealed that this gene was inactivated by transposon insertions or point mutations, indicating that melanin biosynthesis in Aeromonas occurs through the homogentisate pathway. Horizontal gene transfer could have contributed to the adaptation of 34melT to a highly polluted environment, as 13 genomic islands were identified in its genome, some of them containing genes coding for fitness-related traits. Heavy metal resistance genes were also found, along with others associated with oxidative and nitrosative stresses. These characteristics, together with melanin production and the ability to use different substrates, may explain the ability of this microorganism to live in an extremely polluted environment.

Influence of light on lignin-degrading activities of fungal genus Polyporus s. str

Six strains belonging to five species of Polyporus (P. arcularius, P. arcularioides, P. tricholoma, P. cfr. tricholoma, and P. varius), collected from an Atlantic Forest area in Misiones (Argentina), where species usually grow exposed to high temperatures and humidity, were identified by morphological and molecular analyses. P. tricholoma (BAFC 4536) and P. arcularioides (BAFC 4534) were selected by their lignin‐degrading enzyme production, their ability to produce primordial of basidiomes under submerged fermentation, and the decrease in lignin content caused in Poplar wood (up to 29% after 45 days). Among several variables evaluated with a Plackett–Burman design (glucose, copper, vanillic acid and manganese concentration, incubation period, and light incidence), the most important factor affecting laccase and Mn‐peroxidase (MnP) production by both strains, was light incidence. Light induced fruit body development but diminished laccase and MnP production. Moreover, a modified isoenzymatic laccase pattern was observed, showing additional isoenzymes when fungi were cultivated under darkness and differences in optimal temperature. Although the studied strains did not produce high laccase and MnP titers (uppermost detected 4230 and 90 U L−1, respectively), their laccases showed thermal stability and optimal temperature above 70 °C, representing an interesting source in the search of thermo‐tolerant enzymes for biotechnological applications.