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Regulation of tyrosylprotein sulfotransferases activity by sulfotyrosine / 中国医学科学院学报
Acta Academiae Medicinae Sinicae ; (6): 241-245, 2007.
Artículo en Chino | WPRIM | ID: wpr-229995
ABSTRACT
<p><b>OBJECTIVE</b>To investigate the role of sulfated tyrosine in regulating the activity of tyrosylprotein sulfotransferases (TPST) 1 and TPST2.</p><p><b>METHODS</b>Constructs of TPST 1 and TPST2 were amplified by polymerase chain reaction (PCR), then fused into immunoglobulin G1 Fc region. All the variants in which sulfated tyrosines were mutated to phenylalanine were made by the PCR-based Quick Change method and confirmed by sequencing the entire reading frame. Small hairpin RNA (shRNA) constructs-targeting nucleotides 259-275 of TPST1 and nucleotides 73-94 of TPST2 were generated and subcloned into pBluescript. Human embryonic kidney (HEK) 293T cells were transfected with these plasmids. One day later, cells were split one part was labeled with 35S-cysteine and methionine or 35S-Na2SO3 overnight, the second part was used for 125I labeled binding experiment, and the third part was retained for binding and flow cytometry.</p><p><b>RESULTS</b>Tyrosines at position 326 of TPST1 and position 325 of TPST2 were sulfated posttranslationally. Tyrosine sulfation of TPSTs was effectively inhibited by sulfation inhibitors, including specific shRNAs and non-specific NaCIO3. shRNAs reduced the sulfation of C3a receptor and C5a receptor, and partially blocked the binding of these two receptors to their respective ligands.</p><p><b>CONCLUSIONS</b>The activities of TPSTs were regulated by tyrosine sulfation. Inhibition of sulfotyrosine decreases the binding ability of C3a receptor and C5a receptor to their respective ligands.</p>
Asunto(s)
Texto completo: Disponible Índice: WPRIM (Pacífico Occidental) Asunto principal: Unión Proteica / Tirosina / Receptores de Complemento / Transfección / Sulfotransferasas / Complemento C3a / Complemento C5a / Línea Celular / Procesamiento Proteico-Postraduccional / Receptor de Anafilatoxina C5a Límite: Humanos Idioma: Chino Revista: Acta Academiae Medicinae Sinicae Año: 2007 Tipo del documento: Artículo

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Texto completo: Disponible Índice: WPRIM (Pacífico Occidental) Asunto principal: Unión Proteica / Tirosina / Receptores de Complemento / Transfección / Sulfotransferasas / Complemento C3a / Complemento C5a / Línea Celular / Procesamiento Proteico-Postraduccional / Receptor de Anafilatoxina C5a Límite: Humanos Idioma: Chino Revista: Acta Academiae Medicinae Sinicae Año: 2007 Tipo del documento: Artículo