Sulfhydryl groups of rabbit liver arylsulfatase A.
Rabbit liver arylsulfatase A (aryl-sulfate sulfhydrolase, EC 3.1.6.1) monomers of 130 kDa contain two free sulfhydryl groups as determined by spectrophotometric titration using 5,5'-dithiobis(2-nitrobenzoate) and by labeling with the fluorescent probe 5-(iodoacetamidoethyl)aminonaphthalene-1-sulfonic acid. Fluorescence quenching data indicate that the reactive sulfhydryl is present in proximity to one or more tryptophan residues. Chemical modification of the sulfhydryl groups does not alter the distinctive pH-dependent aggregation property of the arylsulfatase A. The free sulfhydryls of the enzyme react with numerous sulfhydryl reagents. Based on the reactions of iodoacetic acid, methyl methanethiosulfonate, 5,5'-dithiobis(2-nitrobenzoate) and 5-(iodoacetamidoethyl)aminonaphthalene-1-sulfonic acid with the sulfhydryl groups of arylsulfatase A, it is concluded that free sulfhydryls are not essential for the enzyme activity. In contrast, the observed inactivation of the enzyme by p-hydroxymercuribenzoate or p-hydroxymercuriphenylsulfonate is probably due to a modification of a histidine residue, consistent with previous reports that histidine is near the active site of arylsulfatase A. p-Hydroxymercuribenzoate and p-hydroxymercuriphenylsulfonate are able to react both with cysteine and with histidine residues of the protein molecule.
Waheed
A
A
Van Etten
R L
RL
D000818:Animals; D002553:Cerebroside-Sulfatase / Q000032:analysis*; D002850:Chromatography, Gel; D004228:Dithionitrobenzoic Acid / Q000494:pharmacology; D005456:Fluorescent Dyes / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D006902:Hydroxymercuribenzoates / Q000494:pharmacology; D008099:Liver / Q000201:enzymology*; D008741:Methyl Methanesulfonate / Q000031:analogs & derivatives / Q000494:pharmacology; D009282:Naphthalenesulfonates / Q000494:pharmacology; D010663:Phenylmercury Compounds / Q000494:pharmacology; D011817:Rabbits; D013050:Spectrometry, Fluorescence; D013429:Sulfatases / Q000032:analysis*; D013438:Sulfhydryl Compounds / Q000032:analysis*; D013997:Time Factors
D005456:Fluorescent Dyes; D006902:Hydroxymercuribenzoates; D009282:Naphthalenesulfonates; D010663:Phenylmercury Compounds; D013438:Sulfhydryl Compounds; C030601:4-hydroxymercuribenzoate; C002910:4-hydroxymercuribenzenesulfonate; C014674:methyl methanethiosulfonate; C006421:1,5-I-AEDANS; D004228:Dithionitrobenzoic Acid; D008741:Methyl Methanesulfonate; D013429:Sulfatases; D002553:Cerebroside-Sulfatase