Journals Information
International Journal of Biochemistry and Biophysics Vol. 10(2), pp. 9 - 23
DOI: 10.13189/ijbb.2022.100201
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Evaluation of Phenol Sulfotransferase (SULT1A1) Ligand Binding Order
Joe D. Beckmann 1,*, Billie Schultz 1, Brian J. Doyle 2
1 Department of Chemistry, Alma College, 614 W. Superior St., Alma, MI, 48801, USA
2 Department of Biology, Alma College, 614 W. Superior St., Alma, MI, 48801, USA
ABSTRACT
Cytosolic sulfotransferases (SULTs) modify and regulate many endogenous and xenobiotic compounds. There are some disagreements regarding the binding pattern of their nucleotide (PAPS, PAP) and acceptor site ligands. Here we present an investigation of the homodimeric bovine phenol sulfotransferase (bSULT1A1) using several methods not dependent on catalytic turnover. Binding order was not determinable from intrinsic protein fluorescence changes in response to PAP(S), 7-hydroxycoumarin (7-HC) or pentachlorophenol (PCP). Equilibrium dialysis showed no binding of 7-HC to apo-bSULT1A1 up to 100 µM ligand, but inclusion of PAP induced biphasic binding of 7-HC. Isothermal titration calorimetry revealed exothermic half-sites binding of PAP to the dimeric protein. No binding of 7-HC to apo-SULT1A1 was detected, but SULT1A1:PAP also bound 7-HC at 0.5 per subunit. Differential scanning fluorimetry failed to show thermal stabilization of protein by excess 7-HC or PCP, whereas inclusion of PAP increased T m dramatically. Susceptibility of bSULT1A1 to cleavage by chymotrypsin was not affected by excess 7-HC, but enzyme with PAP ± 7-HC resisted proteolysis. Excess 7-HC did not change protein cysteine reactivity with DTNB, whereas PAP impeded the fast phase of reaction attributable to C168. These observations indicate an ordered binding scheme for bSULT1A1. Testing other SULT ligand binding patterns with these methods is suggested.
KEYWORDS
Sulfotransferase, SULT1A1, Phenol, 7-Hydroxycoumarin, PAPS, Binding Order, Dialysis, Calorimetry, Kinetics, Proteolysis
Cite This Paper in IEEE or APA Citation Styles
(a). IEEE Format:
[1] Joe D. Beckmann , Billie Schultz , Brian J. Doyle , "Evaluation of Phenol Sulfotransferase (SULT1A1) Ligand Binding Order," International Journal of Biochemistry and Biophysics, Vol. 10, No. 2, pp. 9 - 23, 2022. DOI: 10.13189/ijbb.2022.100201.
(b). APA Format:
Joe D. Beckmann , Billie Schultz , Brian J. Doyle (2022). Evaluation of Phenol Sulfotransferase (SULT1A1) Ligand Binding Order. International Journal of Biochemistry and Biophysics, 10(2), 9 - 23. DOI: 10.13189/ijbb.2022.100201.