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Comparison of extraction methods for recovery of extracellular beta-glucosidase in two different forest soils

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Masciandaro, Grazia, Macci, Cristina, Doni, Serena, Maserti, Bianca Elena, Leo, A. Calvo-Bado, Ceccanti, Brunello and Wellington, E. M. H. (Elizabeth M. H.), 1954-. (2008) Comparison of extraction methods for recovery of extracellular beta-glucosidase in two different forest soils. Soil Biology & Biochemistry, Vol.40 (No.9). pp. 2156-2161. ISSN 0038-0717

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Abstract

A study was made of the efficiency of three different extractants, 0.1 M sodium pyrophosphate (pH 7), 67 mM phosphate buffer (pH 6) and 0.5 M potassium sulphate (pH 6.6), in recovering the protein quantity and the beta-glucosidase enzyme activity from two natural forest soils: (1) an inceptisoil located in Tuscany (Italy) in a mild Mediterranean climate, and (2) a Lithic Calcixeroll soil located in Murcia (southeast of Spain) in a dry-semiarid climate. The pyrophosphate was used to determine the activity of extracellular-humic-bound proteins, while the phosphate buffer and potassium sulphate were used to extract dissolved extracellular proteins. The latter extractant, after chloroform fumigation, was also used to measure total proteins in soil. A preliminary screening, using SDS-PAGE in one dimension, was also carried out in order to optimize the separation condition of soil proteins extracted with different buffers. To remove the interfering co-extracted substances (humic acid) a purification step using a column packed with insoluble polyvinylpyrrolidone was performed. The highest beta-glucosidase activity was recovered in the pyrophosphate extract, thus confirming its capability of extracting humic-bound beta-glucosidase enzyme in a stable and active form. The extractants performed differently with the two soil types and band patterns obtained with SDS-PAGE were extractant-specific, demonstrating that each was selective for a particular class of proteins. Surprisingly, protein bands were also obtained using pyrophosphate, in spite of the very dark extract colour due to the presence of humic substances. (C) 2008 Elsevier Ltd. All rights reserved.

Item Type: Journal Article
Subjects: Q Science > QD Chemistry
S Agriculture > SD Forestry
Divisions: Faculty of Science > Life Sciences (2010- ) > Biological Sciences ( -2010)
Library of Congress Subject Headings (LCSH): Forest soils, Humic acid -- Derivatives, Proteins, Extraction (Chemistry), Glucosidases, Enzymes
Journal or Publication Title: Soil Biology & Biochemistry
Publisher: Elsevier
ISSN: 0038-0717
Official Date: September 2008
Volume: Vol.40
Number: No.9
Number of Pages: 6
Page Range: pp. 2156-2161
Identification Number: 10.1016/j.soilbio.2008.05.001
Status: Peer Reviewed
Publication Status: Published
Access rights to Published version: Restricted or Subscription Access
Funder: European Commission (EC), Consiglio nazionale delle ricerche (Italy) (CNR)
Grant number: 505450 (EC)
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URI: http://wrap.warwick.ac.uk/id/eprint/29317

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