Substrate specificity of human monoamine (M)-form phenol sulfotransferase: preparation and analysis of Dopa 3-O-sulfate and Dopa 4-O-sulfate.
Upon two-dimensional thin-layer separation, the sulfated L-3, 4-dihydroxyphenylalanine (L-DopaS) generated enzymatically was found to co-migrate with only one of the two ninhydrin-stained spots corresponding to the two sulfated forms (3-O-sulfate and 4-O-sulfate) of synthetic L-DopaS. To clarify precisely the identity of the enzymatically generated L-DopaS, the two sulfated forms of synthetic L-DopaS were separated and purified using high performance liquid chromatography. Purified L-Dopa 3-O-sulfate and L-Dopa 4-O-sulfate were identified by 1H-nuclear magnetic resonance (NMR) spectrometry and used as standards in the analysis of the L-DopaS generated during metabolic labeling of HepG2 human hepatoma cells or enzymatic assay using recombinant human monoamine (M)-form phenol sulfotransferase. The results obtained demonstrated unequivocally the generation of L-Dopa 3-O-sulfate, indicating the specificity of the M-form phenol sulfotransferase being for the meta-hydroxyl group of L-Dopa.
Suiko
M
M
Department of Biochemistry, University of Texas Health Center at Tyler, TX 75710, USA.
Sakakibara
Y
Y
Awan-Khan
R
R
Sakaida
H
H
Yoshikawa
H
H
Ranasinghe
J G
JG
Liu
M C
MC
D015239:Arylsulfotransferase / Q000378:metabolism*; D006528:Carcinoma, Hepatocellular; D002855:Chromatography, Thin Layer; D006801:Humans; D007527:Isoenzymes / Q000378:metabolism*; D007980:Levodopa / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism; D008113:Liver Neoplasms; D009682:Magnetic Resonance Spectroscopy; D013379:Substrate Specificity; D013431:Sulfates / Q000378:metabolism; D013462:Sulfur Radioisotopes; D013463:Sulfuric Acid Esters / Q000737:chemistry / Q000378:metabolism*; D014407:Tumor Cells, Cultured
C116412:3,4-dihydroxyphenylalanine 3-O-sulfate; C116413:3,4-dihydroxyphenylalanine 4-O-sulfate; D007527:Isoenzymes; D013431:Sulfates; D013462:Sulfur Radioisotopes; D013463:Sulfuric Acid Esters; D007980:Levodopa; D015239:Arylsulfotransferase