Purification and characterization of an acid phosphatase from Trichoderma harzianum

Abstract

An acid phosphatase from Trichoderma harzianum was purified in a single step using a phenyl-Sepharose chromatography column. A typical procedure showed 22-fold purification with 56% yield. The purified enzyme showed as a single band on SDS-PAGE with an apparent molecular weight of 57.8 kDa. The pH optimum was 4.8 and maximum activity was obtained at 55 degrees C. The enzyme retained 60% of its activity after incubation at 55 degrees C for 60 min. The K (m) and V (max) values for p-nitrophenyl phosphate (p-NPP) as a substrate were 165 nM and 237 nM min(-1), respectively. The enzyme was partially inhibited by inorganic phosphate and strongly inhibited by tungstate. Broad substrate specificity was observed with significant activities for p-NPP, ATP, ADP, AMP, fructose 6-phosphate, glucose 1-phosphate and phenyl phosphate.

Effect of acid phosphatase produced by Trichoderma asperellum Q1 on growth of Arabidopsis under salt stress

“…In this study, the ACPase from T. asperellum Q1 was purified by a combination of ammonium sulphate precipitation, ion-exchange, and gel filtration chromatography. The purified enzyme showed as a single band on SDS-PAGE and the apparent molecular weight was generally consistent with that of T. harzianum (Leitão et al 2010); however, it was obviously less than that from A. caespitosus (186 and 190 kDa) (Guimarães et al 2004) and greater than that from A. fumigatus (18 kDa) (Bernard et al 2002).…”

“…But its role in P-solubilization differs from different kinds of microorganisms. Shen et al (2011) demonstrated that most plant growth-promoting rhizo-bacteria can excrete ACPase, and some studies have also been conducted on the purification and characterization of ACPase from fungi (Shimizu 1993;Guimarães et al 2004;Leitão et al 2010). An ACPase from T. harzianum was purified in a single step using a phenyl-sepharose chromatography column.…”

“…An ACPase from T. harzianum was purified in a single step using a phenyl-sepharose chromatography column. Its molecular weight was approximately 57.8 kDa, and the optimum pH and temperature was 4.8 and 55°C, respectively (Leitão et al 2010). In this study, the ACPase from T. asperellum Q1 was purified by a combination of ammonium sulphate precipitation, ion-exchange, and gel filtration chromatography.…”

Avaliação De Fontes De Fosfato Para Produção De Fosfatase Ácida Em Fermentação Submersa

“…Ensaio enzimático: A atividade da fosfatase ácida foi medida de acordo com (Leitão et al, 2010), utilizando como substrato fosfato de p-nitrofenilo sal disódico hexahidratado (p-NPP) (Sigma Aldrich ™). Uma unidade (1U) de atividade da fosfatase ácida será definida como 1 μM de p-nitrofenol (p-NP) formado por minuto (Ames, 1966).…”

“…Entre os microrganismos solubilizantes de fosfato, as estirpes dos gêneros bacterianos Pseudomonas, Bacillus, Rhizobium e fungos tais como Aspergillus sp., Humicola sp., Mucor sp., Penicillium sp., Metarhizium sp. e Trichoderma harzianum são poderosos solubilizantes de fosfato (Guimarães et al, 2004;Boyce e Walsh, 2007;Leitão, et al 2010).…”