Induction and characterization of  -galactosidase in an extreme thermophile.
A thermostable beta-galactosidase (EC 3.2.1.23; beta-dgalactoside galactohydrolase) was found to be inducible in an extreme thermophile resembling Thermus aquaticus. Enzyme induction was achieved by the addition of lactose, galactose, or the alpha-galactoside, melibiose, to growing cultures. The addition of glucose to induced cultures had a repressive effect on further enzyme synthesis. The enzyme was purified 78-fold, and the optimum temperature and pH for activity were determined to be 80 C and pH 5.0, respectively. The enzyme was activated by both manganese and ferrous iron. Sulfhydryl activation and thermal stabilization indicate that the thermophilic beta-galactosidase is a sulfhydryl enzyme. Kinetic determinations at 80 C established a K(m) of 2.0 x 10(-3)m for the chromogenic substrate o-nitrophenyl beta-d-galactopyranoside (ONPG) and a K(1) of 7.5 x 10(-3)m for lactose. The Arrhenius energy of activation (for the hydrolysis of ONPG) was calculated to be 13.7 kcal/mole. A molecular weight of 5.7 x 10(5) daltons was estimated by elution of the enzyme from Sephadex 4B.
D000096:Acetone; D000645:Ammonium Sulfate; D001419:Bacteria / Q000201:enzymology* / Q000254:growth & development; D002474:Cell-Free System; D011232:Chemical Precipitation; D002850:Chromatography, Gel; D004187:Disaccharides / Q000494:pharmacology; D004789:Enzyme Activation; D004790:Enzyme Induction; D004794:Enzyme Repression; D005690:Galactose / Q000494:pharmacology; D005696:Galactosidases / Q000096:biosynthesis* / Q000302:isolation & purification / Q000378:metabolism; D005947:Glucose / Q000494:pharmacology; D006027:Glycosides / Q000494:pharmacology; D006358:Hot Temperature; D006863:Hydrogen-Ion Concentration; D007501:Iron / Q000494:pharmacology; D007785:Lactose / Q000494:pharmacology; D008345:Manganese / Q000494:pharmacology; D008970:Molecular Weight; D013438:Sulfhydryl Compounds / Q000494:pharmacology; D014871:Water Microbiology
D004187:Disaccharides; D006027:Glycosides; D013438:Sulfhydryl Compounds; D000096:Acetone; D008345:Manganese; D007501:Iron; D005696:Galactosidases; D005947:Glucose; D007785:Lactose; D000645:Ammonium Sulfate; D005690:Galactose