Glycerol protection and purification of Bacillus subtilis glucose dehydrogenase.
Bacillus subtilis glucose dehydrogenase (EC 1.1.1.47) has been purified from sporulating cell extract to apparent homogeneity (as determined by polyacrylamide gel electrophoresis, sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and isoelectric focusing). The enzyme purified as a single molecular species with no evidence for a multiple form of the enzyme. The B. subtilis glucose dehydrogenase has an apparent isoelectric point of 4.7-4.8 and an apparent Mr = 126,000 and is comprised of four subunits of Mr = 31,500 each. The glucose 2-deoxyglucose and glucosamine substrate specificity of the enzyme is similar to the substrate specificity for B. subtilis spore germination, suggesting that the spore glucose dehydrogenase may play some role in spore germination. The B. subtilis glucose dehydrogenase is extremely dependent on the presence of glycerol or other hydrophobic bond-stabilizing agents (or NAD) for retention of enzymatic activity, and the presence of glycerol (20% w/v) in the extraction and purification buffers was absolutely necessary for the successful purification of this enzyme.
D001412:Bacillus subtilis / Q000201:enzymology*; D002237:Carbohydrate Dehydrogenases / Q000378:metabolism*; D004355:Drug Stability; D042843:Glucose 1-Dehydrogenase; D005948:Glucose Dehydrogenases / Q000302:isolation & purification / Q000378:metabolism*; D005990:Glycerol / Q000494:pharmacology*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008970:Molecular Weight; D013171:Spores, Bacterial / Q000201:enzymology
D002237:Carbohydrate Dehydrogenases; D005948:Glucose Dehydrogenases; D042843:Glucose 1-Dehydrogenase; D005990:Glycerol