b-Xylosidases are hydrolytic enzymes which play an important role in xylan degradation, hydrolyzing xylobiose and xylooligosaccharides to xylose from the nonreducing end. Filamentous fungi are particularly interesting producers of this enzyme from an industrial point of view, due to the fact that they secrete b-xylosidases into the medium. Besides, fungal b-xylosidases are highly advantageous for their elevated activity levels and specificity. Interest in xylanolytic enzymes has been increasing, for their possible application in many biotechnological processes. This fact has driven the isolation, purification and characterization of several b-xylosidases. In this review, the mechanisms of action, substrate specificities, physicochemical characteristics, regulation at molecular level, molecular cloning and classification of filamentous fungal b-xylosidases are described. The potential industrial applications of fungal b-xylosidases will also be presented.

Two xylanases from the crude culture filtrate of Penicillium sclerotiorum were purified to homogeneity by a rapid and efficient procedure, using ion-exchange and molecular exclusion chromatography. Molecular masses estimated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis were 23.9 and 33.1 kDa for xylanase I and II, respectively. The native enzymes' molecular masses of 23.8 and 30.8 kDa were estimated for xylanase I and II, respectively, by molecular exclusion chromatography. Both enzymes are glycoproteins with optimum temperature and pH of 50 degrees C and pH 2.5 for xylanase I and 55 degrees C and pH 4.5 for xylanase II. The reducing agents beta-mercaptoethanol and dithio-treitol enhanced xylanase activities, while the ions Hg(2+) and Cu(2+) as well the detergent SDS were strong inhibitors of both enzymes, but xylanase II was stimulated when incubated with Mn(2+). The K (m) value of xylanase I for birchwood xylan and for oat spelt xylan were 6.5 and 2.6 mg mL(-1), respectively, whereas the K (m) values of xylanase II for these substrates were 26.61 and 23.45 mg mL(-1). The hydrolysis of oat spelt xylan by xylanase I released xylobiose and larger xylooligosaccharides while xylooligosaccharides with a decreasing polymerization degree up to xylotriose were observed by the action of xylanase II. The present study is among the first works to examine and describe an extracellular, highly acidophilic xylanase, with an unusual optimum pH at 2.5. Previously, only one work described a xylanase with optimum pH 2.0. This novel xylanase showed interesting characteristics for biotechnological process such as feed and food industries.

In recent decades, xylanases have been used in many processing industries. This study describes the xylanase production by Penicillium glabrum using brewer's spent grain as substrate. Additionally, this is the first work that reports the purification and characterization of a xylanase using this agroindustrial waste. Optimal production was obtained when P. glabrum was grown in liquid medium in pH 5.5, at 25 °C, under stationary condition for six days. The xylanase from P. glabrum was purified to homogeneity by a rapid and inexpensive procedure, using ammonium sulfate fractionation and molecular exclusion chromatography. SDS-PAGE analysis revealed one band with estimated molecular mass of 18.36 kDa. The optimum activity was observed at 60 °C, in pH 3.0. The enzyme was very stable at 50 °C, and high pH stability was verified from pH 2.5 to 5.0. The ion Mn2+ and the reducing agents β-mercaptoethanol and DTT enhanced xylanase activity, while the ions Hg2+, Zn2+, and Cu2+ as well as the detergent SDS were strong inhibitors of the enzyme. The use of brewer's spent grain as substrate for xylanase production cannot only add value and decrease the amount of this waste but also reduce the xylanase production cost.

In recent decades, beta-xylosidases have been used in many processing industries. In this work, the study of xylosidase production by Penicillium sclerotiorum and its characterization are reported. Optimal production was obtained in medium supplemented with oat spelts xylan, pH 5.0, at 30 degrees C, under stationary condition for six days. The optimum activity temperature was 60 degrees C and unusual optimum pH 2.5. The enzyme was stable at 50 and 55 degrees C, with half-life of 240 and 232min, respectively. High pH stability was verified from pH 2.0 to 4.0 and 7.5. The beta-xylosidase was strongly inhibited by divalent cations, sensitive to denaturing agents SDS, EDTA and activated by thiol-containing reducing agents. The apparent V(max) and K(m) values was 0.48micromol PNXPmin(-1)mg(-1) protein and 0.75mM, respectively. The enzyme was xylose tolerant with a K(i) of 28.7. This enzyme presented interesting characteristics for biotechnological process such as animal feed, juice and wine industries.

ResumoMuitas enzimas produzidas por fungos têm relevantes aplicações em diferentes áreas industriais. O objetivo deste trabalho foi selecionar fungos filamentosos produtores de enzimas hidrolíticas, bem como estabelecer os resíduos agroindustriais capazes de induzir maiores níveis de produção. As atividades xilanásica, endoglucanásica, amilásica e poligalacturonásica foram determinadas incubandose os filtrados de cultura com seus respectivos substratos. Posteriormente, a determinação de açúcares redutores foi realizada utilizando-se o reagente ácido 3,5-dinitrosalicílico. A determinação de proteínas foi realizada segundo o método de Bradford modificado. Dentre as linhagens fúngicas avaliadas, Aspergillus niger J4 apresentou maiores níveis de produção de xilanases (8,73 ± 0,34 U/mL) e esta foi maior quando o bagaço de malte foi utilizado como substrato (9,80 ± 0,02 U/mL). Penicillium miczynskii produziu os índices mais elevados de atividade endoglucanásica (0,13 ± 0,03 U/mL), sendo está última favorecida na presença de casca de abacaxi (0,18 ± 0,02 U/mL). Em relação à amilase, A. niger J26 foi selecionada como a melhor linhagem produtora (6,10 ± 0,30 U/mL), sendo o farelo de trigo estabelecido como o melhor substrato indutor de sua produção (7,32 ± 0,14 U/mL). Penicillium verruculosum exibiu os maiores níveis de atividade poligalacturonásica (8,65 ± 0,12 U/mL), especialmente quando cultivado em presença de casca de laranja (10,32 ± 0,10 U/mL). O emprego destes resíduos no processo de produção destas enzimas poderá não apenas reduzir seus custos de produção, como também diminuir, substancialmente, o impacto ambiental causado pela deposição destes resíduos no ambiente. Palavras-chave: Linhagens fúngicas, hidrolases, substratos lignocelulósicos, amiláceos, substâncias pécticas AbstractMany fungal enzymes have relevant applications in different industrial areas. The objective of this work was to select fungi producing hydrolytic enzymes, as well as establish agro-industrial wastes capable of inducing higher production levels. Xylanase, endoglucanase, amylase and poligalacturonase activities were determined by incubating the culture filtrates with their respective substrates. Subsequently, the reducing sugars determination was carried out using 3,5-dinitrosalicylic acid reagent. The protein determination was performed according the modified Bradford method. Among the fungal strains evaluated, Aspergillus niger J4 showed higher levels of xylanase production (8.73 ± 0.34 U/ml) and this