Reversible covalent inhibition of a phenol sulfotransferase by coenzyme A.
Phenol sulfotransferases (SULTs), which normally bind 3'-phosphoadenosine-5'-phosphosulfate as the donor substrate, are inhibited by CoA and its thioesters. Here, we report that inhibition of bovine SULT1A1 by CoA is time-dependent at neutral pH under non-reducing conditions. The rates of inactivation by CoA indicate an initial reversible SULT:CoA complex with a dissociation constant of 5.7 microM and an inactivation rate constant of 0.07 min(-1). Titrations with CoA and prolonged incubations reveal that inactivation of the dimeric enzyme is stoichiometric, consistent with the observation of complete conversion of the protein to a slightly decreased electrophoretic mobility. Both activity and normal electrophoretic migration are restored by 2-mercaptoethanol. Mutagenesis demonstrated that Cys168 is the site of CoA adduction, and a consistent model was constructed that reveals a new SULT molecular dynamic. Cysteine reaction kinetics with Ellman's reagent revealed a PAPS-induced structural change consistent with the model that accounts for binding of CoA.
Chodavarapu
Sundari
S
Department of Biochemistry, Alma College, Alma, MI 48801, USA.
Hertema
Heather
H
Huynh
Tien
T
Odette
Jessica
J
Miller
Rachel
R
Fullerton
Aaron
A
Alkirwi
Jason
J
Hartsfield
D'Juan
D
Padmanabhan
Kaillathe
K
Woods
Caleb
C
Beckmann
Joe D
JD
D000818:Animals; D015239:Arylsulfotransferase / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000235:genetics; D002417:Cattle; D003065:Coenzyme A / Q000737:chemistry*; D003545:Cysteine / Q000737:chemistry / Q000235:genetics; D004789:Enzyme Activation; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008623:Mercaptoethanol / Q000737:chemistry; D008958:Models, Molecular; D009154:Mutation; D013440:Sulfides / Q000737:chemistry*
D013440:Sulfides; D008623:Mercaptoethanol; D015239:Arylsulfotransferase; D003545:Cysteine; D003065:Coenzyme A