Analysis of the substrate specificity of human sulfotransferases SULT1A1 and SULT1A3: Site-directed mutagenesis and kinetic studies

Brix, Lulu A., Barnett, Amanda C., Duggleby, Ronald G., Leggett, Barbara and McManus, Michael E. (1999) Analysis of the substrate specificity of human sulfotransferases SULT1A1 and SULT1A3: Site-directed mutagenesis and kinetic studies. Biochemistry, 38 32: 10474-10479.


Author Brix, Lulu A.
Barnett, Amanda C.
Duggleby, Ronald G.
Leggett, Barbara
McManus, Michael E.
Title Analysis of the substrate specificity of human sulfotransferases SULT1A1 and SULT1A3: Site-directed mutagenesis and kinetic studies
Journal name Biochemistry   Check publisher's open access policy
Publication date 1999
Sub-type Article
DOI 10.1021/bi990795q
Volume number 38
Issue number 32
ISSN 0006-2960
Start page 10474
End page 10479
Total pages 6
Editor Gordon G. Hammes
Place of publication Washington, D.C.
Publisher American Chemical Society
Collection year 1999
Language eng
Subject C1
270108 Enzymes
780105 Biological sciences
Abstract Sulfonation is an important metabolic process involved in the excretion and in some cases activation of various endogenous compounds and xenobiotics. This reaction is catalyzed by a family of enzymes named sulfotransferases. The cytosolic human sulfotransferases SULT1A1 and SULT1A3 have overlapping yet distinct substrate specificities. SULT1A1 favors simple phenolic substrates such as p-nitrophenol, whereas SULT1A3 prefers monoamine substrates such as dopamine. In this study we have used a variety of phenolic substrates to functionally characterize the role of the amino acid at position 146 in SULT1A1 and SULT1A3. First, the mutation A146E in SULT1A1 yielded a SULT1A3-like protein with respect to the Michaelis constant for simple phenols. The mutation E146A in SULT1A3 resulted in a SULT1A1-like protein with respect to the Michaelis constant for both simple phenols and monoamine compounds. When comparing the specificity of SULT1A3 toward tyramine with that for p-ethylphenol (which differs from tyramine in having no amine group on the carbon side chain), we saw a 200-fold preference for tyramine. The kinetic data obtained with the E146A mutant of SULT1A3 for these two substrates clearly showed that this protein preferred substrates without an amine group attached. Second, changing the glutamic acid at position 146 of SULT1A3 to a glutamine, thereby neutralizing the negative charge at this position, resulted in a 360-fold decrease in the specificity constant for dopamine. The results provide strong evidence that residue 146 is crucial in determining the substrate specificity of both SULT1A1 and SULT1A3 and suggest that there is a direct interaction between glutamic acid 146 in SULT1A3 and monoamine substrates.
Keyword Biochemistry & Molecular Biology
Sulfation
Cdnas
Biology
Q-Index Code C1

Document type: Journal Article
Sub-type: Article
Collection: School of Chemistry and Molecular Biosciences
 
Versions
Version Filter Type
Citation counts: TR Web of Science Citation Count  Cited 42 times in Thomson Reuters Web of Science Article | Citations
Google Scholar Search Google Scholar
Access Statistics: 67 Abstract Views  -  Detailed Statistics
Created: Mon, 13 Aug 2007, 11:18:41 EST