Exploring the compatibility between hydrothermal depolymerization of alkaline lignin from sugarcane bagasse and metabolization of the aromatics by bacteria.

Although hydrothermal treatments for biomass fractionation have been vastly studied, their effect on the depolymerization of isolated lignins in terms of yield, composition, and compatibility of the produced lignin bio-oils with bioconversion is still poorly investigated. In this study, we evaluated the hydrothermal depolymerization of an ß-O-4'-rich lignin extracted from sugarcane bagasse by alkaline fractionation, investigating the influence of temperature (200-350 °C), time (30-90 min), and solid-liquid ratio (1:10-1:50 m.v-1) on yield of bio-oils (up to 31 wt%) rich in monomers (light bio-oils). Principal Components Analysis showed that the defunctionalization of the aromatic monomers was more pronounced in the most severe reaction conditions and that the abundance of more hydrophobic monomers increased in more diluted reactions. While the high-molecular-weight (heavy) bio-oil generated at 350 °C, 90 min, and 1:50 m.v-1 failed to support bacterial growth, the corresponding light bio-oil rich in aromatic monomers promoted the growth of bacteria from 9 distinct species. The isolates Pseudomonas sp. LIM05 and Burkholderia sp. LIM09 showed the best growth performance and tolerance to lignin-derived aromatics, being the most promising for the future development of biological upgrading strategies tailored for this lignin stream.

Correction to: Two Novel Acetylesterases from Pantoea dispersa: Recombinant Expression, Purification, and Characterization.

The original version of this article unfortunately contained a mistake. Under Materials and Methods heading, Bacterial Strains sub-heading, the correct name of the used strain is "FEI4 65" and not "FzEI4 65."

Production of Galactose Oxidase Inside the Fusarium fujikuroi Species Complex and Recombinant Expression and Characterization of the Galactose Oxidase GaoA Protein from Fusarium subglutinans.

Galactose oxidase catalyzes a two-electron oxidation, mainly from the C6 hydroxyl group of D-galactose, with the concomitant reduction of water to hydrogen peroxide. This enzyme is secreted by Fusarium species and has several biotechnological applications. In this study, a screening of galactose oxidase production among species of the Fusarium fujikuroi species complex demonstrated Fusarium subglutinans to be the main producer. The truncated F. subglutinans gaoA gene coding for the mature galactose oxidase was expressed from the prokaryotic vector pTrcHis2B in the E. coli Rosetta™ (DE3) strain. The purified recombinant enzyme presented temperature and pH optima of 30 °C and 7.0, respectively, KM of 132.6 ± 18.18 mM, Vmax of 3.2 ± 0.18 µmol of H2O2/min, kcat of 12,243 s-1, and a catalytic efficiency (kcat/KM) of 9.2 × 104 M-1 s-1. In the presence of 50% glycerol, the enzyme showed a T50 of 59.77 °C and was stable for several hours at pH 8.0 and 4 °C. Besides D-(+)-galactose, the purified enzyme also acted against D-(+)-raffinose, α-D-(+)-melibiose, and methyl-α-D-galactopyranoside, and was strongly inhibited by SDS. Although the F. subglutinans gaoA gene was successfully expressed in E. coli, its endogenous transcription was not confirmed by RT-PCR.

Two Novel Acetylesterases from Pantoea dispersa: Recombinant Expression, Purification, and Characterization.

Two novel acetylesterases from Pantoea dispersa, with low amino acid sequence identity between them, were expressed in Escherichia coli with a carboxyl-His6 tail given by the expression plasmid, purified, and characterized. The purified proteins, named Est-1 and Est-2, had a molecular mass of 33 kDa and 37 kDa, respectively. Both proteins presented a modeled structure of homodimers with monomers presenting the α/ß-hydrolase fold, with the catalytic triad Ser-Asp-His present in the active site. The KM for p-nitrophenyl acetate and Vmax values found for Est-1 were of 1.4 ± 0.2 mM and 8.66 ± 0.59 µmol/min and for Est-2 were of 0.36 ± 0.077 mM and 6.13 ± 0.56 µmol/min, respectively. Both enzymes presented an optimum pH of 7.0. The optimum temperature for Est-1 was 40 °C and for Est-2 was 50 °C. The temperatures in which the enzymes Est-1 and Est-2 lost half of their activity (T50) were 44.1 and 58.9 °C, respectively. SDS, EDTA, and PMSF significantly inhibited the enzymes. The two purified enzymes also presented activity against triacetin and were able to deacetylate the carbohydrates pectin and xylan, with higher activity against pectin. Thus, they could be considered as carbohydrate esterases.