Beta-D-phosphogalactoside galactohydrolase from Streptococcus cremoris HP: purification and enzyme properties.
beta-d-phosphogalactoside galactohydrolase (beta-PGal) was isolated and purified from cell-free extracts of Streptococcus cremoris HP to apparent homogeneity to gel electrophoresis. Using the chromogenic o-nitrophenol-beta-d-galactopyranoside-6-phosphate as substrate, the purified enzyme exhibited a specific activity of 18.71 U/mg of protein and K(m) and V(max) values of 5.88 x 10(-4) M and 23.8 mumol of o-nitrophenol liberated per min per mg of protein, respectively. d-Galactose-6-phosphate was a weak competitive inhibitor of beta-PGal. Activity was relatively heat resistant and was maximal from pH 5.0 to 8.0 and over a temperature range of 45 to 52 C. Dithiothreitol, ethylenediaminetetraacetic acid, and citrate stimulated beta-PGal activity, whereas Mg(2+), Li(1+), and p-hydroxymercuribenzoate were inhibitory. Molecular weight of the enzyme was estimated at 6.76 x 10(4). Amino acid composition was similar to other beta-phosphogalactosidases previously investigated, with the exception that the S. cremoris enzyme contains a small amount of half cystine.
D000596:Amino Acids / Q000032:analysis; D002474:Cell-Free System; D002848:Chromatography, DEAE-Cellulose; D002850:Chromatography, Gel; D002951:Citrates / Q000494:pharmacology; D004229:Dithiothreitol / Q000494:pharmacology; D004492:Edetic Acid / Q000494:pharmacology; D005033:Ethylmaleimide / Q000494:pharmacology; D005690:Galactose / Q000378:metabolism; D005696:Galactosidases / Q000032:analysis / Q000302:isolation & purification* / Q000378:metabolism; D006027:Glycosides / Q000378:metabolism; D006600:Hexosephosphates / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D006902:Hydroxymercuribenzoates / Q000494:pharmacology; D007461:Iodoacetates / Q000494:pharmacology; D008094:Lithium / Q000494:pharmacology; D008274:Magnesium / Q000494:pharmacology; D009578:Nitrobenzenes / Q000378:metabolism; D009596:Nitrophenols / Q000096:biosynthesis; D010710:Phosphates / Q000378:metabolism; D013291:Streptococcus / Q000201:enzymology*; D013696:Temperature
D000596:Amino Acids; D002951:Citrates; D006027:Glycosides; D006600:Hexosephosphates; D006902:Hydroxymercuribenzoates; D007461:Iodoacetates; D009578:Nitrobenzenes; D009596:Nitrophenols; D010710:Phosphates; D008094:Lithium; D004492:Edetic Acid; D005696:Galactosidases; D008274:Magnesium; D005033:Ethylmaleimide; D004229:Dithiothreitol; D005690:Galactose