Interaction of pantetheinase with sulfhydryl reagents and disulfides.
The effect of many thiol reagents and disulfides on pantetheinase (E.C. 3.5.1.-; pantetheine hydrolase) was studied in the presence or absence of S-pantetheine-3-pyruvate as substrate. Iodoacetamide, iodoacetate, bromopyruvate and N-ethylmaleimide irreversibly inactivate the enzyme at very different rates. Inactivation constants, corrected for the different reactivity of halogeno derivatives with non-protein thiols, suggest the presence of an essential sulfhydryl group in the enzyme and a negatively charged environment near this group. p-Chloromercuribenzoate is the most effective inhibitor; 2-nitro-5-thiocyanobenzoate, o-iodosobenzoate and hydrogen peroxide give a biphasic inhibition pattern, indicating the existence of two sulfhydryl groups whose modification affects activity. Organic arsenicals decrease activity to about 50%. Neutral and positively charged disulfides are effective inhibitors. Substrate protects the enzyme from inactivation, except in the case of negatively charged disulfides, where the presence of substrate enhances the inhibitory effect. Titration with Ellman's reagent or 4,4'-dithiodipyridine under various experimental conditions demonstrated the existence of two sulfhydryls and three disulfides in the fully active enzyme. Pantetheinase may become inactive during purification with concomitant loss of one titrable sulfhydryl group.
D000477:Alkylating Agents / Q000494:pharmacology; D000581:Amidohydrolases / Q000037:antagonists & inhibitors* / Q000378:metabolism; D004220:Disulfides / Q000494:pharmacology*; D058851:GPI-Linked Proteins; D007700:Kinetics; D010204:Pantetheine / Q000031:analogs & derivatives / Q000378:metabolism; D013438:Sulfhydryl Compounds / Q000378:metabolism; D013439:Sulfhydryl Reagents / Q000494:pharmacology*
D000477:Alkylating Agents; D004220:Disulfides; D058851:GPI-Linked Proteins; D013438:Sulfhydryl Compounds; D013439:Sulfhydryl Reagents; D010204:Pantetheine; C043983:pantetheine-3-pyruvate; D000581:Amidohydrolases; C022314:pantetheinase