Crystallization and preliminary crystallographic analysis of GanB, a GH42 intracellular β-galactosidase from Geobacillus stearothermophilus

Hodaya V. Solomon, Orly Tabachnikov, Hadar Feinberg, Lata Govada, Naomi E. Chayen, Yuval Shoham*, Gil Shoham

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

19 Scopus citations

Abstract

Geobacillus stearothermophilus T-6 is a Gram-positive thermophilic soil bacterium that contains a multi-enzyme system for the utilization of plant cell-wall polysaccharides, including xylan, arabinan and galactan. The bacterium uses a number of endo-acting extracellular enzymes that break down the high-molecular-weight polysaccharides into decorated oligosaccharides. These oligosaccharides enter the cell and are further hydrolyzed into sugar monomers by a set of intracellular glycoside hydrolases. One of these intracellular degrading enzymes is GanB, a glycoside hydrolase family 42 β-galactosidase capable of hydrolyzing short β-1,4-galactosaccharides to galactose. GanB and related enzymes therefore play an important part in the hemicellulolytic utilization system of many microorganisms which use plant biomass for growth. The interest in the biochemical characterization and structural analysis of these enzymes stems from their potential biotechnological applications. GanB from G. stearothermophilus T-6 has recently been cloned, overexpressed, purified, biochemically characterized and crystallized in our laboratory as part of its complete structure-function study. The best crystals obtained for this enzyme belong to the primitive orthorhombic space group P212121, with average crystallographic unit-cell parameters of a = 71.84, b = 181.35, c = 196.57 Å. Full diffraction data sets to 2.45 and 2.50 Å resolution have been collected for both the wild-type enzyme and its E323A nucleophile catalytic mutant, respectively, as measured from flash-cooled crystals at 100 K using synchrotron radiation. These data are currently being used for the full three-dimensional crystal structure determination of GanB.

Original languageAmerican English
Pages (from-to)1114-1119
Number of pages6
JournalActa Crystallographica Section F: Structural Biology and Crystallization Communications
Volume69
Issue number10
DOIs
StatePublished - Oct 2013

Keywords

  • GH42
  • GanB
  • Geobacillus stearothermophilus
  • β-galactosidase

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