ABSTRACT
alpha-Glucosidase from two microbial sources, Bacillus stearothermophilus and Brewer's yeast, has been used to catalyze transglycosylation reactions and a comparative study was carried out to determine the regioselectivity of this reaction. Bacterial alpha-glucosidase exhibited higher transfer activity with maltose and was able to synthesize tri- and tetrasaccharides in high yield (27%). In the case of yeast enzyme, only trisaccharides were synthesized in lower yield. Structure analysis of transglycosylation products by means of GC-MS and NMR spectroscopy revealed a correlation between the hydrolytic substrate specificity and the regioselectivity of transglycosylation reaction. Higher substrate specificity of bacterial enzyme, however, influenced its transglucosylation activity toward other saccharide acceptors.
Subject(s)
Oligosaccharides/chemical synthesis , alpha-Glucosidases/metabolism , Chromatography, High Pressure Liquid , Gas Chromatography-Mass Spectrometry , Glycosylation , Hydrolysis , Magnetic Resonance Spectroscopy , Stereoisomerism , Substrate SpecificityABSTRACT
A cell-free extract of a morphologically unstable strain of Dipodascus magnusii contained six proteins with activity of glucose-6-phosphate dehydrogenase (G6PDH). Two of these proteins displayed only NADP(+)-dependent activity, two could utilize both NAD+ and NADP+, but had higher activity with NAD+, and two possessed only NAD(+)-dependent activity. When the cultivation was carried out in the presence of monoiodoacetic acid, only two proteins with G6PDH activity were produced, one of them NAD(+)-dependent and the other NADP(+)-dependent. In all cases, NAD(+)-dependent activity was less stable in the presence of proteinases than was the NADP(+)-dependent activity.
Subject(s)
Fungal Proteins/isolation & purification , Glucosephosphate Dehydrogenase/isolation & purification , Isoenzymes/isolation & purification , NADP/metabolism , NAD/metabolism , Saccharomycetales/enzymology , Fungal Proteins/metabolism , Glucosephosphate Dehydrogenase/metabolism , Isoenzymes/metabolismABSTRACT
alpha-Glucosidase is an enzyme widely used in biochemical analytical methods. Aspergillus niger was selected as a potential source for its production. Conditions for glucosidase production were optimized and the enzyme was isolated from the culture supernatant by dialysis and anion-exchange chromatography. The activity of the enzyme was determined by maltose hydrolysis to glucose, which was determined using a glucose-specific electrode or by high-performance liquid chromatography. The isolated enzyme was further characterized by sodium dodecyl sulphate-polyacrylamide gel electrophoresis, substrate specificity and fast protein liquid chromatography. The Michaelis constant, optimal temperature and stability of the enzyme preparation were determined.
Subject(s)
Aspergillus niger/enzymology , alpha-Glucosidases/isolation & purification , Anions , Catalase/metabolism , Chromatography, Ion Exchange , Electrodes , Enzymes, Immobilized , Glucose/metabolism , Glucose Oxidase/metabolism , Substrate Specificity , Ultrafiltration , alpha-Glucosidases/metabolismSubject(s)
Aldose-Ketose Isomerases , Candida/enzymology , Peptide Hydrolases/isolation & purification , Streptomyces/enzymology , Carbohydrate Epimerases/isolation & purification , Caseins/analysis , Protein Denaturation , Proteins/analysis , Urate Oxidase/isolation & purification , Uric Acid/analysisABSTRACT
The papers of Kolhouse et al. and Cooper et al. described the occurrence of vitamin B12 analogues of unknown origin in blood serum. Some of these analogues may be derived from slaughter cattle raised on feed supplemented with vitamins and minerals, as was observed by Allen. Herbert et al. found vitamin B12 analogues in multivitamin preparations produced in U.S.A., and Kanazawa et al. in human liver, red cells and brain. It is not clear so far, if and how do vitamin B12 analogues interfere with vitamin B12 metabolism. When P. freudenreichii was cultivated in the presence of o-phenylenediamine and 5,6-dimethylbenzimidazole, which may be considered precursors as well as antimetabolites of vitamin B12, the stimulation of biosynthesis of substances with biological activity of vitamin B12 took place. Various signs show that these substances are probably vitamin B12 analogues. During stimulated and nonstimulated production of vitamin B12 by P. freudenreichii, two substances with vitamin B12 biological activity have always been obtained. Their relation was not stable and differed according to the conditions of cultivation. Every attempt to stimulate the biosynthesis of vitamin B12 resulted in the suppression of production of the substance with higher molecular weight, even if the biosynthesis of cobalamin (lower molecular weight) was increased. In our note we want to pay attention to the character of substances arising in the stimulated biosynthesis.
