Este estudo teve como objetivo confirmar, estatisticamente, atraves de experimentos controlados a... more Este estudo teve como objetivo confirmar, estatisticamente, atraves de experimentos controlados a contribuicao de Caiman crocodilus yacare para a ciclagem de nutrientes na lagoa Baia das Pedras, Pantanal de Pocone - MT. Estudos com este enfoque foram iniciados no Pantanal apenas no final da decada de 90, na lagoa Baia das Pedras, onde, durante o periodo de estiagem, foram contabilizados cerca de 1000 individuos da especie Caiman crocodilus yacare em apenas 20.800 m2, area da lagoa. Nesse periodo de estiagem a produtividade da lagoa e muito elevada em relacao ao periodo da cheia, quando a agua fica transparente, sendo que os jacares desempenham naquele periodo um importante papel. Foi utilizado para o experimento um cativeiro de 8x6 m, com uma unica caida para agua. No centro do cativeiro ha um tanque de concreto de 4x2 m e com 30 cm de profundidade. O animal foi colocado apos a estabilizacao das variaveis limnologicas do tanque. Foram coletadas 6 amostras de agua, sendo tres antes e...
Multienzymatic complexes with plant lignocellulose-degrading activities have recently been identi... more Multienzymatic complexes with plant lignocellulose-degrading activities have recently been identified in filamentous fungi secretomes. Such complexes have potential biotechnological applications in the degradation of agro-industrial residues. Fungal species from the Clonostachys genus have been intensively investigated as biocontrol agents; however so far their use as producers of lignocellulose-degrading enzymes has not been extensively explored. Secretomes of Clonostachys byssicola following growth on different carbon sources (passion fruit peel, soybean hulls, cotton gin trash, banana stalk, sugarcane bagasse, orange peel, and a composition of soybean hulls: cotton gin trash:orange peel) were subjected to enzymatic assays. Remarkable differences were observed among the samples, especially regarding levels of mannanase and pectinase activities. Secretomes were then subjected to Blue Native PAGE in order to resolve putative protein complexes which subsequently had their composition...
ABSTRACT The fungus Trichoderma reesei is a major producer of plant cell wall-degrading enzymes, ... more ABSTRACT The fungus Trichoderma reesei is a major producer of plant cell wall-degrading enzymes, such as cellulases and hemicellulases. These enzymes have been used to hydrolyze agro-industrial wastes in processes of second-generation bio-fuel production. The disaccharide lactose, the main by-product of dairy industry, is considered the most attractive substrate for the commercial production of cellulases by T. reesei. In the present work, it was shown that T. reesei, cultivated in media supplemented with either lactose or galac-tose, secrete proteins which behave as multi-enzymatic complexes. Blue Native electrophoresis (BN-PAGE) displayed two major bands which were specific or present in higher amounts in each secretome sample, complex I in galactose medium and complex II in lactose medium. The protein components of complexes I and II were identified by means of LC-MS/MS. The complexes were composed of different glycosyl hydrolases and predicted proteins related to biopolymer metabolism. Zymography coupled to BN-PAGE showed that T. reesei secretome presents higher cellulolytic and xylanolytic activities in lactose than in galactose-induced medium. The results presented here corroborate the existence of hydrolysis based on multi-enzymatic complexes in T. reesei secretome, as observed previously in other fungi, such as Trichoderma harzianum and Penicillium purpurogenum.
Acta crystallographica. Section F, Structural biology and crystallization communications, 2008
A binary complex of the Schizolobium parahyba chymotrypsin inhibitor (SPCI) with chymotrypsin was... more A binary complex of the Schizolobium parahyba chymotrypsin inhibitor (SPCI) with chymotrypsin was purified by size-exclusion chromatography and crystallized by the sitting-drop vapour-diffusion method with 100 mM MES-NaOH pH 5.5, 20%(w/v) PEG 6000, 200 mM LiCl as precipitant and 200 mM nondetergent sulfobetaine molecular weight 201 Da (NDSB-201) as an additive. SPCI is a small protein with 180 amino-acid residues isolated from S. parahyba seeds and is able to inhibit chymotrypsin at a 1:1 molar ratio by forming a stable complex. X-ray data were collected to 2.8 A resolution from a single crystal of the SPCI-chymotrypsin binary complex under cryogenic conditions. The crystal belongs to space group P2(1)2(1)2(1), with unit-cell parameters a = 45.28, b = 64.57, c = 169.23 A, and the R(merge) is 0.122 for 11 254 unique reflections. A molecular-replacement solution was found using the preliminary crystal structure of SPCI and the structure of chymotrypsin (PDB code 4cha) independently as...
The major royal jelly protein 1 (MRJP1) is the main glycoprotein in honey bee royal jelly. In bra... more The major royal jelly protein 1 (MRJP1) is the main glycoprotein in honey bee royal jelly. In brain tissues, MRJP1 is found in intercellular spaces and associated to cytoskeleton within cells. MRJP1 must be involved in multiple biological functions, yet there is a lack of structural information on the protein. MRJP1 was herein purified from royal jelly and characterized through electrophoresis and mass spectrometry as the same protein found in cerebral tissue. Unfolding curves obtained by circular dichroism analyses strongly suggest its high stability under different pHs. However, calcium ions made MRJP1 susceptible to temperature and pH effects. In the presence of 2 mM calcium, very high stabilities were achieved at pH 6.0 and 7.0 with ΔG 25 over 62 kJ mol−1. Overall, the present results represent a valuable effort aimed at the structural characterization of MRJP1, representing an essential step toward the determination of its roles in honey bee neural processes.
Este estudo teve como objetivo confirmar, estatisticamente, atraves de experimentos controlados a... more Este estudo teve como objetivo confirmar, estatisticamente, atraves de experimentos controlados a contribuicao de Caiman crocodilus yacare para a ciclagem de nutrientes na lagoa Baia das Pedras, Pantanal de Pocone - MT. Estudos com este enfoque foram iniciados no Pantanal apenas no final da decada de 90, na lagoa Baia das Pedras, onde, durante o periodo de estiagem, foram contabilizados cerca de 1000 individuos da especie Caiman crocodilus yacare em apenas 20.800 m2, area da lagoa. Nesse periodo de estiagem a produtividade da lagoa e muito elevada em relacao ao periodo da cheia, quando a agua fica transparente, sendo que os jacares desempenham naquele periodo um importante papel. Foi utilizado para o experimento um cativeiro de 8x6 m, com uma unica caida para agua. No centro do cativeiro ha um tanque de concreto de 4x2 m e com 30 cm de profundidade. O animal foi colocado apos a estabilizacao das variaveis limnologicas do tanque. Foram coletadas 6 amostras de agua, sendo tres antes e...
Multienzymatic complexes with plant lignocellulose-degrading activities have recently been identi... more Multienzymatic complexes with plant lignocellulose-degrading activities have recently been identified in filamentous fungi secretomes. Such complexes have potential biotechnological applications in the degradation of agro-industrial residues. Fungal species from the Clonostachys genus have been intensively investigated as biocontrol agents; however so far their use as producers of lignocellulose-degrading enzymes has not been extensively explored. Secretomes of Clonostachys byssicola following growth on different carbon sources (passion fruit peel, soybean hulls, cotton gin trash, banana stalk, sugarcane bagasse, orange peel, and a composition of soybean hulls: cotton gin trash:orange peel) were subjected to enzymatic assays. Remarkable differences were observed among the samples, especially regarding levels of mannanase and pectinase activities. Secretomes were then subjected to Blue Native PAGE in order to resolve putative protein complexes which subsequently had their composition...
ABSTRACT The fungus Trichoderma reesei is a major producer of plant cell wall-degrading enzymes, ... more ABSTRACT The fungus Trichoderma reesei is a major producer of plant cell wall-degrading enzymes, such as cellulases and hemicellulases. These enzymes have been used to hydrolyze agro-industrial wastes in processes of second-generation bio-fuel production. The disaccharide lactose, the main by-product of dairy industry, is considered the most attractive substrate for the commercial production of cellulases by T. reesei. In the present work, it was shown that T. reesei, cultivated in media supplemented with either lactose or galac-tose, secrete proteins which behave as multi-enzymatic complexes. Blue Native electrophoresis (BN-PAGE) displayed two major bands which were specific or present in higher amounts in each secretome sample, complex I in galactose medium and complex II in lactose medium. The protein components of complexes I and II were identified by means of LC-MS/MS. The complexes were composed of different glycosyl hydrolases and predicted proteins related to biopolymer metabolism. Zymography coupled to BN-PAGE showed that T. reesei secretome presents higher cellulolytic and xylanolytic activities in lactose than in galactose-induced medium. The results presented here corroborate the existence of hydrolysis based on multi-enzymatic complexes in T. reesei secretome, as observed previously in other fungi, such as Trichoderma harzianum and Penicillium purpurogenum.
Acta crystallographica. Section F, Structural biology and crystallization communications, 2008
A binary complex of the Schizolobium parahyba chymotrypsin inhibitor (SPCI) with chymotrypsin was... more A binary complex of the Schizolobium parahyba chymotrypsin inhibitor (SPCI) with chymotrypsin was purified by size-exclusion chromatography and crystallized by the sitting-drop vapour-diffusion method with 100 mM MES-NaOH pH 5.5, 20%(w/v) PEG 6000, 200 mM LiCl as precipitant and 200 mM nondetergent sulfobetaine molecular weight 201 Da (NDSB-201) as an additive. SPCI is a small protein with 180 amino-acid residues isolated from S. parahyba seeds and is able to inhibit chymotrypsin at a 1:1 molar ratio by forming a stable complex. X-ray data were collected to 2.8 A resolution from a single crystal of the SPCI-chymotrypsin binary complex under cryogenic conditions. The crystal belongs to space group P2(1)2(1)2(1), with unit-cell parameters a = 45.28, b = 64.57, c = 169.23 A, and the R(merge) is 0.122 for 11 254 unique reflections. A molecular-replacement solution was found using the preliminary crystal structure of SPCI and the structure of chymotrypsin (PDB code 4cha) independently as...
The major royal jelly protein 1 (MRJP1) is the main glycoprotein in honey bee royal jelly. In bra... more The major royal jelly protein 1 (MRJP1) is the main glycoprotein in honey bee royal jelly. In brain tissues, MRJP1 is found in intercellular spaces and associated to cytoskeleton within cells. MRJP1 must be involved in multiple biological functions, yet there is a lack of structural information on the protein. MRJP1 was herein purified from royal jelly and characterized through electrophoresis and mass spectrometry as the same protein found in cerebral tissue. Unfolding curves obtained by circular dichroism analyses strongly suggest its high stability under different pHs. However, calcium ions made MRJP1 susceptible to temperature and pH effects. In the presence of 2 mM calcium, very high stabilities were achieved at pH 6.0 and 7.0 with ΔG 25 over 62 kJ mol−1. Overall, the present results represent a valuable effort aimed at the structural characterization of MRJP1, representing an essential step toward the determination of its roles in honey bee neural processes.
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