ABSTRACT
The xylanase (Xyn10B) that strongly adsorbs on microcrystalline cellulose was isolated from Driselase. The Xyn10B contains a Carbohydrate-binding module family 1 (CBM1) (IrpCBMXyn10B) at N-terminus. The canonical essential aromatic residues required for cellulose binding were conserved in IrpCBMXyn10B; however, its adsorption ability was markedly higher than that typically observed for the CBM1 of an endoglucanase from Trametes hirsuta (ThCBMEG1). An analysis of the CBM-GFP fusion proteins revealed that the binding capacity to cellulose (7.8 µmol/g) and distribution coefficient (2.0 L/µmol) of IrpCBMXyn10B-GFP were twofold higher than those of ThCBMEG1-GFP (3.4 µmol/g and 1.2 L/µmol, respectively), used as a reference structure. Besides the canonical aromatic residues (W24-Y50-Y51) of typical CBM1-containing proteins, IrpCBMXyn10B had an additional aromatic residue (Y52). The mutation of Y52 to Ser (IrpCBMY52S-GFP) reduced these adsorption parameters to 4.4 µmol/g and 1.5 L/µmol, which were similar to those of ThCBMEG1-GFP. These results indicate that Y52 plays a crucial role in strong cellulose binding.
Subject(s)
Basidiomycota/enzymology , Endo-1,4-beta Xylanases/chemistry , Endo-1,4-beta Xylanases/metabolism , Tyrosine/metabolism , Amino Acid Sequence , Binding Sites , Cellulose/chemistry , Cellulose/metabolism , Chemical Fractionation , Cotton Fiber , Endo-1,4-beta Xylanases/genetics , Fungal Proteins/chemistry , Fungal Proteins/metabolism , Glycoside Hydrolases/chemistry , Glycoside Hydrolases/metabolism , Green Fluorescent Proteins/genetics , Green Fluorescent Proteins/metabolism , Molecular Sequence Data , Mutation , Recombinant Fusion Proteins/genetics , Recombinant Fusion Proteins/metabolismABSTRACT
In order to provide healthy experimental animals, it is important to find and remove animals that have been accidentally exposed to various stresses during breeding. This study focuses mouse health-care management. Here we used human olfaction and gas chromatography-mass spectrometry to assess odor intensity and determine the concentrations of odor components. The feces were collected from mice that were exposed to 4 different stresses (no bedding chips, shaking, fasting, and movement restriction). These stresses caused a change in odor intensity as assessed by 6 panelists. Seventeen components were identified as dominant components in the odor that was emitted from feces. The concentration of each compound was converted to relative values versus its odor threshold levels in order to select ones effective for the quality of the odor. As a result, 12 selected components were found to be a useful set for the recognition of mice bred under different stress conditions. The present results may provide useful information for the development of standard fecal odor materials that may be used for the training of mouse care personnel.
