ABSTRACT
Activity of α-L-arabinofuranosidae (EC 3.2.1.55) ranged between 0.03 and 1.63 U/ml whenSclerotium rolfsii was grown in a synthetic medium containing different nitrogen and carbon sources. Agricultural and forest residues such as rice straw, corn cob, groundnut husk, and apple pomace were suitable substrates for enzyme production. Maximum activity, 4.2 U/ml, was with pre-treated corn cobs.
ABSTRACT
Simultaneous Saccharification and Fermentation (SSF) experiments were carried out on agricultural residues using culture filtrate of Sclerotium rolfsii, which produces high levels of cellulases and hemicellulases for the saccharification of rice straw and bagasse, and Candida shehatae--the D-xylose fermenting yeast, and Saccharomyces cerevisiae, both separately and in coculture, for fermenting the released sugars. The coculture system showed efficient utilization of hydrolyzed sugars with 30-38% and 10-13% increase in ethanol production as compared to C. shehatae and S. cerevisiae, respectively, when cultivated separately. SSF simulation studies were carried out using standard sugar mixtures of glucose, xylose, and cellobiose. Both organisms could not use cellobiose, whereas glucose was used preferentially. C. shehatae was capable of utilizing xylose in the presence of glucose.
Subject(s)
Candida/metabolism , Carbohydrate Metabolism , Saccharomyces cerevisiae/metabolism , Xylose/metabolism , Candida/enzymology , Cellobiose/metabolism , Ethanol/metabolism , Fermentation , Glucose/metabolism , Oryza , Saccharomyces cerevisiae/enzymologyABSTRACT
A gene encoding exo-1,4-beta-D-glucosidase, from Ruminococcus albus AR67, was cloned in Escherichia coli, restriction mapped, and shown to be expressed from sequences within the insert that function as a promoter in E. coli. The cloned enzyme was located predominantly in the cytoplasm (40%) and attached to insoluble cell components (48%). After purification to homogeneity, the enzyme (Mr = 64,000, monomeric) was specific for substrates with beta-D-glucopyranosyl configuration and was inactive against alpha-glucosides, lactosides and xylosides. Km values of the enzyme decreased with increasing chain length (G2-G5). Glucose was the major product of hydrolysis from cellodextrins. Preference for longer chain cellodextrins is consistent with exo-1,4-beta-D-glucan glucohydrolase mode of action [E.C. 3.2.1.74].
Subject(s)
Glucosidases/metabolism , Peptococcaceae/enzymology , beta-Glucosidase/metabolism , Chromatography, Gel , Chromatography, Ion Exchange , Cloning, Molecular , Electrophoresis, Polyacrylamide Gel , Gene Library , Glucan 1,4-beta-Glucosidase , Kinetics , Peptococcaceae/genetics , Plasmids , Recombinant Proteins/isolation & purification , Recombinant Proteins/metabolism , Restriction Mapping , Subcellular Fractions/enzymology , Substrate Specificity , Ultrafiltration , beta-Glucosidase/genetics , beta-Glucosidase/isolation & purificationABSTRACT
Pennisetin, the alcohol soluble storage protein of pearl millet (Pennisetum americanum), was isolated in a homogeneous state. The intrinsic viscosity [n] of this protein was found to be in the range of 16.5-17.7 ml/g in 70% (v/v) aqueous ethanol. The [eta] changed marginally when temperature was increased from 20 to 70 degrees C and also in the presence of 10 mM NaCl. The data indicated that pennisetin was a rigid, rod shaped asymmetric hydrodynamic particle with molecular dimensions in the range of 301 x 14.4 A - 317.7 x 14.2 A. During denaturation with guanidine hydrochloride (Gdn.HCl), the intrinsic viscosity of pennisetin increased from 16 to 25ml/g with a mid point at 3.6 M of the denaturant. The native protein structure was unfolded in 6 M Gdn.HCl as shown by the exposure of aromatic amino acid residues buried in the native state and this transition was found to be reversible. The intrinsic viscosity of pennisetin in 5.9 M Gdn.HCl corresponded to Mr 25,000 which was comparable to that determined by SDS-PAGE.
Subject(s)
Edible Grain/chemistry , Panicum/chemistry , Plant Proteins , Electrophoresis, Polyacrylamide Gel , Molecular Weight , Plant Proteins/isolation & purification , Prolamins , Protein Conformation , Spectrometry, Fluorescence , ViscosityABSTRACT
The endoglucanase component (CMCase I) ofPenicillium funiculosum cellulase was purified to apparent homogeneity by ultrafiltration and gel chromatography. It consists of a single polypeptide chain with a molecular weight of 56000 and is a glycoprotein. Viscometric and end-product analysis revealed the randomness of enzyme action. Multifunctional characteristic of CMCase I was studied with various carbohydrate substrates.
