A cytosolic trehalase from the thermophilic fungus Humicola grisea var. thermoidea

Cardello, Leonardo; Terenzi, Héctor Francisco; Jorge, João Atı́lio

doi:10.1099/13500872-140-7-1671

Cited by 29 publications

(25 citation statements)

References 20 publications

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“…In addition, the pH optimum of approx. 4.0 (Thevelein, 1984) and the apparent K m value of 0.38 mM (in comparison with 0.29-4.7 mM found by Londesborough & Varimo (1984), Mittenbu$ hler & Holzer (1988), Sumida et al (1989) and Cardello et al (1994)) are comparable to those of other enzymes obtained from fungi.…”

Section: supporting

confidence: 64%

Identification and characterization of an extracellular acid trehalase from the ectomycorrhizal fungus Amanita muscaria

et al. 2000

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The non-reducing disaccharide trehalose is a major storage compound in most fungi. For a better understanding of carbon metabolism in the ectomycorrhizal-forming basidiomycete Amanita muscaria, trehalase activity was analysed from mycelium growing in liquid culture or on agar plates, and from mycorrhizas. Trehalase activity was found in both the culture medium and in mycelial extracts. The excreted trehalase was purified to apparent homogeneity by anion-exchange chromatography, gel filtration and preparative gel electrophoresis. The identified enzyme belongs to the group of acid trehalases. The apparent molecular mass of the native enzyme was estimated to be 165 kDa by gel filtration and 135 kDa by SDS-PAGE. Isoelectrofocusing indicated an isoelectric point (pI) of approx. 3.7, which is typical for acid trehalases. The enzyme is highly specific for its substrate trehalose, with an apparent K m of 0.38 mM. Validamycin and sucrose act as competitive inhibitors with K i values of 45 nM and 15 mM, respectively. The activity of acid trehalase excreted into the growth medium is independent of the carbon source (glucose or trehalose), revealing that the enzyme is not regulated by its substrate trehalose. Nevertheless, fungal hyphae grown in the absence of an external carbon source showed enhanced enzyme excretion. The biochemical characteristics of the trehalase activity measured in the mycelium are in the same range as those determined for the excreted enzyme. The enzyme is localized in the cell wall and its activity is strongly decreased in ectomycorrhizas.

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Section: supporting

confidence: 64%

Identification and characterization of an extracellular acid trehalase from the ectomycorrhizal fungus Amanita muscaria

et al. 2000

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“…The mycelial enzyme (M r 360 kDa) and the conidial enzyme (M r 580 kDa) of H. grisea var. thermoidea contained 12% and 56% carbohydrate, respectively [20,21]. The mycelial and the secreted trehalases of S. thermophilum contained 51% and 82% carbohydrate, respectively, but had similar molecular mass (M r 370 kDa) [22].…”

Section: Puri¢cation Of Trehalasesmentioning

confidence: 99%

A comparison of thermal characteristics and kinetic parameters of trehalases from a thermophilic and a mesophilic fungus

Bharadwaj

1999

FEMS Microbiology Letters

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Trehalases from a thermophilic fungus Thermomyces lanuginosus (M r 145 kDa) and a mesophilic fungus Neurospora crassa (M r 437 kDa) were purified to compare their thermal characteristics and kinetic constants. Both trehalases were maximally active at 50³C, had an acidic pH optimum and were glycoproteins (20% and 43%, w/w, carbohydrate content for T. lanuginosus and N. crassa, respectively). At their temperature optimum, their K m was similar (0.57 and 0.52 mM trehalose, for T. lanuginosus and N. crassa, respectively) but the V max of N. crassa enzyme was nine times higher than of T. lanuginosus enzyme. The catalytic efficiency, k cat /K m , for N. crassa trehalase was one order of magnitude higher (6.2U10 6 M 31 s 31 ) than of T. lanuginosus trehalase (4U10 5 M 31 s 31 ). At their T opt (50³C), trehalase from both sources exhibited similar thermostability (t 1a2 s 6 h). The energy of activation, E a , for T. lanuginosus trehalase was 15.12 kcal mol 31 and for N. crassa trehalase it was 9.62 kcal mol 31 . The activation energy for thermal inactivation for the N. crassa enzyme (92 kcal mol 31 ) was two-fold higher than for the T. lanuginosus enzyme (46 kcal mol 31 ). The present study shows that the trehalase of N. crassa is not only more stable but also a better catalyst than the T. lanuginosus enzyme. ß

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“…Usually, high production of trehalases by thermophilic fungi occurs in media containing mannitol (Cardello et al, 1994) or starch (Almeida et al, 1999), but not when the molds are grown in glucose as a carbon source. Here, high levels of trehalase production by aged cultures of Malbranchea were observed using glucose as the main carbon source, although the enzyme was detected in the culture medium only after a complete consumption of glucose (data not shown).…”

Section: Effect Of Potential Inhibitors and Activators On Trehalase Amentioning

confidence: 99%

“…sulfurea, produced and secreted into the culture medium in the presence of starch as the main carbon source. Although sharing properties of acid and neutral trehalases, the extracellular Malbranchea enzyme is inhibited by Ca 2+ and insensitive to ATP, in contrast to other thermophilic fungi trehalases (Cardello et al, 1994;Kadowaki et al, 1996;Almeida et al, 1999), and, amongst other interesting properties, is also insensitive to Ag + . The enzyme seems to be controlled by catabolic regulatory circuits and to be involved in the assimilation of exogenous trehalose as a source of carbon.…”

mentioning

confidence: 91%

Biochemical properties of an extracellular trehalase from Malbranchea pulchella var. Sulfurea

et al. 2011

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The thermophilic fungus Malbranchea pulchella var. sulfurea produced good amounts of extracellular trehalase activity when grown for long periods on starch, maltose or glucose as the main carbon source. Studies with young cultures suggested that the main role of the extracellular acid trehalase is utilizing trehalose as a carbon source. The specific activity of the purified enzyme in the presence of manganese (680 U/mg protein) was comparable to that of other thermophilic fungi enzymes, but many times higher than the values reported for trehalases from other microbial sources. The apparent molecular mass of the native enzyme was estimated to be 104 kDa by gel filtration and 52 kDa by SDS-PAGE, suggesting that the enzyme was composed by two subunits. The carbohydrate content of the purified enzyme was estimated to be 19 % and the pi was 3.5. The optimum pH and temperature were 5.0-5.5 and 55° C, respectively. The purified enzyme was stimulated by manganese and inhibited by calcium ions, and insensitive to ATP and ADP, and 1 mM silver ions. The apparent K(M) values for trehalose hydrolysis by the purified enzyme in the absence and presence of manganese chloride were 2.70 ± 0.29 and 2.58 ± 0.13 mM, respectively. Manganese ions affected only the apparent V(max), increasing the catalytic efficiency value by 9.2-fold. The results reported herein indicate that Malbranchea pulchella produces a trehalase with mixed biochemical properties, different from the conventional acid and neutral enzymes and also from trehalases from other thermophilic fungi.

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A cytosolic trehalase from the thermophilic fungus Humicola grisea var. thermoidea

Cited by 29 publications

References 20 publications

Identification and characterization of an extracellular acid trehalase from the ectomycorrhizal fungus Amanita muscaria

Identification and characterization of an extracellular acid trehalase from the ectomycorrhizal fungus Amanita muscaria

A comparison of thermal characteristics and kinetic parameters of trehalases from a thermophilic and a mesophilic fungus

Biochemical properties of an extracellular trehalase from Malbranchea pulchella var. Sulfurea

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