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Measurement of the Galactanase Activity of the GanB Galactanase Protein from Bacillus subtilis   

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Original research article

A brief version of this protocol appeared in:
Journal of Bacteriology
Oct 2016

Abstract

The activity of the endo-β-1,4-galactanase GanB from B. subtilis on the high molecular weight β-1,4-galactan was determined quantitatively by the measurement of the increase of the reducing power or with the dyed substrate Azo-galactan. The generated degradation products were analyzed using thinlayer-chromatography (TLC) or high-performance anion-exchange chromatography (HPAEC).

Keywords: Galactanase-assay, Endo-β-galactanase, Galactan, AZCL-galactan, GanB from Bacillus subtilis, Galacto-oligosaccharides

Background

Bacillus subtilis possesses comprehensive systems for the utilization of plant cell wall polysaccharides including the gene cluster ganSPQAB which encodes galactan utilization elements (Watzlawick et al., 2016). Galactans are high molecular weight galacto-saccharides and are found as side chains of rhamnogalacturonan type I in pectin. Its degradation is carried out by endo-beta1,4 galactanases (EC 3.2.1.89). The utilization of galactan by B. subtilis involves the extracellular galactanase GanB, cleaving the high molecular galactan inside the chain and resulting short oligomers that enter the cell wall to get there further degraded. The ganB gene from B. subtilis was cloned and expressed in E.coli (Watzlawick et al., 2016) and the enzymatic properties of the purified His-tagged mature protein were characterized by galactanase assays.

Copyright: © 2017 The Authors; exclusive licensee Bio-protocol LLC.
How to cite: Watzlawick, H. (2017). Measurement of the Galactanase Activity of the GanB Galactanase Protein from Bacillus subtilis. Bio-protocol 7(7): e2206. DOI: 10.21769/BioProtoc.2206.
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