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.

Searching for monooxygenases and hydrolases in bacteria from an extreme environment.

Microbial oxidation potentials of extremophiles recovered from Pampo Sul oil field, Campos Basin, Brazil, in pure culture or in consortia, were investigated using high-throughput screening (HTS) and multibioreactions. Camphor (1), cis-jasmone (2), 2-methyl-cyclohexanone (3), 1,2-epoxyoctane (4), phenylethyl acetate (5), phenylethyl propionate (6), and phenylethyl octanoate (7) were used to perform multibioreaction assays. Eighty-two bacterial isolates were recovered from oil and formation water samples and those presenting outstanding activities in HTS assays were identified by sequencing their 16S rRNA genes. These results revealed that most microorganisms belonged to the genus Bacillus and presented alcohol dehydrogenase, monooxygenase, epoxide hydrolase, esterase, and lipase activities.

Influence of sugarcane bagasse variability on sugar recovery for cellulosic ethanol production.

In the context of cellulosic ethanol production, special attention must be given to the raw material, as it affects final product yield. As observed for sugarcane, bagasse variations may derive from several elements, for instance edaphoclimatic factors, seasonality, maturation stage and harvesting techniques. Therefore, in the present work, to investigate the impact of raw material characteristics on process performance, sugarcane bagasse from four harvests from October/2010 to October/2011 was pretreated by steam explosion and had its soluble and insoluble solids contents measured, following enzymatic hydrolysis to assess glucan conversion. As confirmed by ANOVA, glucose concentration was related to the solids content in the reactor, whereas glucan conversion was related to the enzymatic load. Variations in raw material composition were indeed observed to significantly interfere in the final sugar recovery, probably due to the increase in the impurities observed as a result of the type of harvest performed in 2011.

Oligomerization as a strategy for cold adaptation: Structure and dynamics of the GH1 ß-glucosidase from Exiguobacterium antarcticum B7.

Psychrophilic enzymes evolved from a plethora of structural scaffolds via multiple molecular pathways. Elucidating their adaptive strategies is instrumental to understand how life can thrive in cold ecosystems and to tailor enzymes for biotechnological applications at low temperatures. In this work, we used X-ray crystallography, in solution studies and molecular dynamics simulations to reveal the structural basis for cold adaptation of the GH1 ß-glucosidase from Exiguobacterium antarcticum B7. We discovered that the selective pressure of low temperatures favored mutations that redesigned the protein surface, reduced the number of salt bridges, exposed more hydrophobic regions to the solvent and gave rise to a tetrameric arrangement not found in mesophilic and thermophilic homologues. As a result, some solvent-exposed regions became more flexible in the cold-adapted tetramer, likely contributing to enhance enzymatic activity at cold environments. The tetramer stabilizes the native conformation of the enzyme, leading to a 10-fold higher activity compared to the disassembled monomers. According to phylogenetic analysis, diverse adaptive strategies to cold environments emerged in the GH1 family, being tetramerization an alternative, not a rule. These findings reveal a novel strategy for enzyme cold adaptation and provide a framework for the semi-rational engineering of ß-glucosidases aiming at cold industrial processes.

A novel cold-adapted and glucose-tolerant GH1 ß-glucosidase from Exiguobacterium antarcticum B7.

A novel GH1 ß-glucosidase (EaBgl1A) from a bacterium isolated from Antarctica soil samples was recombinantly overexpressed in Escherichia coli cells and characterized. The enzyme showed unusual pH dependence with maximum activity at neutral pH and retention of high catalytic activity in the pH range 6 to 9, indicating a catalytic machinery compatible with alkaline conditions. EaBgl1A is also a cold-adapted enzyme, exhibiting activity in the temperature range from 10 to 40°C with optimal activity at 30°C, which allows its application in industrial processes using low temperatures. Kinetic characterization revealed an enzymatic turnover (Kcat) of 6.92s(-1) (cellobiose) and 32.98s(-1) (pNPG) and a high tolerance for product inhibition, which is an extremely desirable feature for biotechnological purposes. Interestingly, the enzyme was stimulated by up to 200 mM glucose, whereas the commercial cocktails tested were found fully inhibited at this concentration. These properties indicate EaBgl1A as a promising biocatalyst for biotechnological applications where low temperatures are required.

Draft Genome Sequence of the Brazilian Toxic Bloom-Forming Cyanobacterium Microcystis aeruginosa Strain SPC777.

Microcystis aeruginosa strain SPC777 is an important toxin-producing cyanobacterium, isolated from a water bloom of the Billings reservoir (São Paulo State, Brazil). Here, we report the draft genome sequence and initial findings from a preliminary analysis of strain SPC777, including several gene clusters involved in nonribosomal and ribosomal synthesis of secondary metabolites.