Aims:  To obtain recombinant strains of Penicillium griseoroseum that produce high levels of pectin lyase (PL) and polygalacturonase (PG) simultaneously. Methods and Results:  A strain with high production of PL was transformed with the plasmid pAN52pgg2, containing the gene encoding PG of P. griseoroseum, under control of the gpd promoter gene from Aspergillus nidulans. Southern blot analysis demonstrated that all strain had at least one copy of pAN52pgg2 integrated into the genome. The recombinant strain P. griseoroseum T20 produced levels of PL and PG that were 266‐ and 27‐fold greater, respectively, than the wild‐type strain. Furthermore, the extracellular protein profile of recombinant T20 showed two protein bands of c. 36 and 38 kDa, associated with PL and PG, respectively. Conclusions:  This recombinant strain T20 produces PL and PG using carbon sources of low costs, and an enzyme preparation that is free of cellulolytic and proteolytic activities. Significance and Impact of the Study:  PL and PG play an important role in the degradation of pectin. Owing to their use in the juice and wines industries, there is a growing interest in the inexpensive production of these enzymes. This work describes an efficient system of protein expression and secretion using the fungus P. griseoroseum.

A novel lectin was isolated from the seeds of Chenopodium quinoa. To achieve this end, the crude extract from the quinoa was submitted to two purification steps, Sephadex G50 and Mono Q. The hemagglutinating activity showed that this lectin agglutinates human erythrocytes. Its activity is inhibited by glucose and mannose, and remained stable under a wide range of pH levels and temperatures. The quinoa lectin was found to be a heterodimeric lectin of approximately 60 kDa, consisting of two subunits of approximately 25 kDa and 35 kDa. This lectin had its antimicrobial activity tested against several bacteria strains and effectively inhibited three strains. These strains were all Gram-negative, making this lectin a promising antimicrobial tool.Keywords: antimicrobial; Chenopodium quinoa; glucose/mannose-specific; lectin; seeds. Practical Application:The purification and characterization of this novel lectin allow its study for a diversity of applications such as antibacterial, anticancer, anti-inflammatory and biotechnology and its further use as drugs for human diseases.

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A novel trypsin inhibitor of protease (CqTI) was purified from Chenopodium quinoa seeds. The optimal extracting solvent was 0.1M NaCl pH 6.8 (p < 0.05). The extraction time of 5h and 90 °C was optimum for the recovery of the trypsin inhibitor from C. quinoa seeds. The purification occurred in gel-filtration and reverse phase chromatography. CqTI presented active against commercial bovine trypsin and chymotrypsin and had a specific activity of 5,033.00 (TIU/mg), which was purified to 333.5-fold. The extent of purification was determined by SDS-PAGE. CqTI had an apparent molecular weight of approximately 12KDa and two bands in reduced conditions as determined by Tricine-SDS-PAGE. MALDI-TOF showed two peaks in 4,246.5 and 7,908.18m/z. CqTI presented high levels of essential amino acids. N-terminal amino acid sequence of this protein did not show similarity to any known protease inhibitor. Its activity was stable over a pH range (2-12), temperatures range (20-100 °C) and reducing agents.Keywords: purification; characterization; inhibitor of trypsin; Chenopodium quinoa; seeds. Practical Application:The purification and characterization of this novel trypsin inhibitor allow its study for a diversity of applications such as antibacterial, anticancer, anti-inflammatory and biotechnology and its further use as drugs for human diseases.

Yeasts capable of growth on xylose were isolated from macaúba (Acrocomia aculeata) fruit, a Brazilian palm tree with great potential for use as biodiesel feedstock production. Candida boidinii UFMG14 strain achieved the highest ethanol production (5 g/L) and was chosen to ferment macaúba presscake hemicellulosic hydrolysate (MPHH). The MPHH was produced by the first time in this work and the resultant fivefold concentrate showed considerable sugar content (52.3 and 34.2 g/L xylose and glucose, respectively) and low furfural (0.01 g/L) and hydroxymethylfurfural (0.15 g/L) concentrations. C. boidinii UFMG14 fermentation was evaluated in supplemented and non-supplemented MPHH containing either 10 or 25 g/L of xylose. The maximum ethanol production (12 g/L) was observed after 48 h of fermentation. The ethanol yield was significantly affected by supplementation and concentration of MPHH while ethanol productivity was affected only by MPHH concentration. This is the first study demonstrating theC. boidinii potential for ethanol production from hemicellulose byproducts.

SCREENING OF SPECIES FROM THE GENUS Penicillium PRODUCING CELL BOUND LIPASES TO BE APPLIED IN THE VEGETABLE OIL HYDROLYSIS. Ten species ofPenicillium genus isolated from different habitats were evaluated as mycelium bound lipase producers to be used in the hydrolysis of vegetable oils. Using olive oil as an inducer three species (P. italicum AT4421, P. janthinellum CCT3162 and P. purpurogenum AT2008) were able to produce lipases having high mycelium bound activities (>150 U g -1 ) and were further characterized in relation to their biochemical and kinetic properties and specificity using vegetable oils having majority fatty acids composition in C12:0 (coconut); C16:0 (palm); C18:1 (canola) and C18:1 (soybean). All the three lipases could enrich the medium with fatty acids according to their respective selectivity and the reaction hydrolysis was found to enhance at least three folds under ultrasonic irradiations. For P. purpurogenum lipase the highest hydrolysis degree (66.8 ± 0.2%) was attained with coconut oil. Both P. italicum and P. janthinellum lipases showed high selectivity for canola oil, resulting in hydrolysis degrees of 79.9 ± 0.5% and 63.5 ± 0.6%, respectively. Analysis of the hydrolysates confirmed that the majority of the fatty acids released by P. italicum and P. janthinellum lipases was composed by oleic acid, and P. purpurogenum lipase the hydrolysate contained approximately 50% of lauric acid.Keywords: lipases; Penicillium; whole-cells; hydrolysis; vegetable oils. INTRODUÇÃODesde sua identificação em 1930, as lipases ainda despertam grande interesse em diversos segmentos industriais. Segundo Daiha et al., 1 há uma projeção de crescimento anual na demanda mundial por estas enzimas estimada em 6,2%, atingindo US$ 345 milhões em 2017. Este aumento na aplicação industrial de lipases tem motivado pesquisas para o isolamento e triagem de novas fontes microbianas desta enzima.As lipases (glicerol éster hidrolases EC 3.1.1.3) fazem parte da classe das hidrolases que atuam em ligações éster-carboxílicas e não requerem cofator. A função biológica primordial destas enzimas é catalisar a hidrólise de triacilgliceróis, entretanto, em condições em que a disponibilidade de água no meio é reduzida, a maioria é capaz de catalisar reações reversas como esterificação e transesterificação (interesterificação, alcoólise e acidólise), entre outras. 2,3 Essa versatilidade e adaptabilidade em diferentes solventes tem resultado no uso difundido das lipases em diversas aplicações industriais, 4,5 incluindo formulação de detergentes, fabricação de aditivos alimentares, controle de pitch na indústria de papel e biocatalisadores para transformação de óleos e gorduras da indústria oleoquímica, como hidrólise de óleos vegetais. 2 A hidrólise enzimática de óleos vegetais é uma alternativa ao processo químico que busca superar os inconvenientes da rota convencional (elevadas condições de temperatura 250 o C e pressão 4,83 mPa), visando fornecer produtos com baixo custo energético por meio da condução da reação em condições mais...

Protoplast fusion between complementary auxotrophic and morphological mutant strains of Penicillium griseoroseum and P. expansum was induced by polyethylene glycol and calcium ions (Ca 2+ ). Fusant strains were obtained in minimal medium and a prototrophic strain, possibly diploid, was chosen for haplodization with the fungicide benomyl. Different recombinant strains were isolated and characterized for occurrence of auxotrophic mutations and pectinolytic enzyme production. The fusant prototrophic did not present higher pectinase production than the parental strains, but among 29 recombinants analyzed, four presented enhanced enzyme activities. The recombinant RGE27, which possesses the same auxotrophic and morphologic mutations as the P. griseoroseum parental strain, presented a considerable increase in polygalacturonase (3-fold) and pectin lyase production (1.2-fold).

Chenopodium quinoa seeds have high protein content. The nutritional value of quinoa is superior compared with traditional cereals. Its essential amino acid composition is considered next to the ideal, and its quality matches that of milk proteins. In this study, the seed storage proteins from Chenopodium quinoa were extracted, fractionated, partially purified, and characterized. The structural characterization was performed by Tricine-SDS-PAGE and two-dimensional electrophoresis, and it confirmed the presence of proteins of molecular weight of 30 and 7kDa, probably corresponding to lectins and trypsin inhibitors, respectively. The functional characterization of these proteins evidenced their activity as antinutritional factors due to their in vitro digestibility. Quinoa proteins have an excellent amino acid composition with many essential amino acids. In vitro digestibility evaluation indicated that heat-treated samples showed a more complete digestion than the native state samples. Quinoa seeds can be an important cereal in human diet after adequate heat treatment.Keywords: Chenopodium quinoa; seeds; lectins; protease inhibitor; in vitro digestibility.Practical Application: SDS-PAGE to evaluate the effect of digestive enzymes in seed proteins.