Galactose-1-phosphatase in rat brain.
A prominent galactose-1-phosphatase was isolated from rat brain and partially purified by chromatography on diethylaminoethyl-Sephacel, hydroxylapatite, and Sephacryl S-300 columns. The galactose-1-phosphatase was separated from alkaline phosphatase, and from two forms of glucose-1-phosphatase. The three columns gave a 10-fold increase in specific activity to 290 mol/min/mg of protein, with a yield of 15%. Of the eight sugar phosphates tested, galactose-1-phosphate was the best substrate for the purified enzyme, followed by glucose-1-phosphate, which was hydrolyzed 40% as rapidly as galactose-1-phosphate. Galactose-1-phosphatase had an optimum pH of 8.5 and a Km value of 2.5 mM for galactose-1-phosphate hydrolysis. Mg2+ was required for activity, and supported half-maximal activity at a concentration of 1.25 mM. Phosphate was the only potent inhibitor found ATP, arsenate, and vanadate caused moderate inhibition of 10 mM levels, whereas AMP, L-homoarginine, and L-phenylalanine stimulated enzyme activity. Galactose-1-phosphatase was determined to have a Stokes radius of 30 A and a sedimentation coefficient of 4.1S. These values were used to calculate a molecular weight of 50,200 and a frictional ratio showing the enzyme to be a globular protein. It is hypothesized that a similar phosphatase may play a role in reducing brain galactose-1-phosphate concentrations in patients with galactosemia.
Gulavita
S J
SJ
Department of Biochemistry, University of Rhode Island, Kingston 02881.
Zhang
L P
LP
Dougherty
J J
JJ
Dain
J A
JA
D000818:Animals; D001921:Brain / Q000201:enzymology*; D002845:Chromatography; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D017886:Durapatite; D005695:Galactosephosphates / Q000302:isolation & purification* / Q000378:metabolism; D006882:Hydroxyapatites; D007700:Kinetics; D008970:Molecular Weight; D010744:Phosphoric Monoester Hydrolases / Q000302:isolation & purification*; D011487:Protein Conformation; D051381:Rats; D011919:Rats, Inbred Strains; D013379:Substrate Specificity
D005695:Galactosephosphates; D006882:Hydroxyapatites; C029973:galactose-1-phosphate; D017886:Durapatite; C069910:galactose-1-phosphatase; D010744:Phosphoric Monoester Hydrolases