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Caracterización de la epóxido hidrolasa SgcF que apoya un intermediario (R) -vicinal diol para la biosíntesis del
Shuangjun Lin1, Geoffrey P Horsman, Yihua Chen
1Division of Pharmaceutical Sciences, University of Wisconsin-Madison, Madison, Wisconsin 53705-2222, USA.
Journal of the American Chemical Society
|October 28, 2009
Resumen
La enzima SgcF es crucial para producir el antibiótico antitumoral C-1027 mediante el procesamiento de un intermediario de epoxido. Esta epoxida hidrolasa genera un diol específico, lo que confirma su papel esencial en la vía de biosíntesis C-1027.
Área de la Ciencia:
- La bioquímica es la bioquímica.
- Biología Molecular Biología Molecular
- Enzimología Enzimología.
Sus antecedentes:
- C-1027 es un antibiótico antitumoral de cromoproteína compuesto por una apoproteína y un cromóforo.
- El cromóforo C-1027 contiene un núcleo de enediína, un aminosugar desoxígeno, un benzoxazolinato y un derivado de la beta-tirosina.
- Se propone una estrategia de biosíntesis convergente para ensamblar el cromóforo C-1027.
Objetivo del estudio:
- Para caracterizar la hidrolasa epóxido SgcF del grupo de genes biosintéticos C-1027.
- Para aclarar el papel de SgcF en la biosíntesis del cromóforo C-1027.
- Investigar la región y la estereospecificidad de la actividad de SgcF en imitaciones de óxido de estireno.
Principales métodos:
- Caracterización in vitro de la enzima SgcF.
- Ensayos enzimáticos que utilizan el óxido de estireno como sustrato.
- Análisis de la producción de C-1027 en una cepa mutante DeltasgcF de Streptomyces globisporus SB1010.
Principales resultados:
- SgcF cataliza la hidrólisis regio- y estereospecífica del óxido de estireno para formar un diol. vicinal.
- La cepa mutante DeltasgcF mostró una producción abolida de C-1027, lo que confirma el papel esencial de sgcF.
- SgcF hidroliza preferentemente el óxido de (S) estireno a (R) fenil diol vicinal con una alta eficiencia (Km = 0,6 mM, kcat = 48 min-1).
- SgcF también hidroliza el óxido de (R) estireno a (R) fenil diol vicinal, pero de manera menos eficiente (Km = 2,0 mM, kcat = 4,3 min-1).
Conclusiones:
- El SgcF juega un papel indispensable en la biosíntesis del C-1027.
- Se identifica un diol (R) -vicinal como un intermediario clave en la vía del cromóforo C-1027.
- La hidrólisis enantioconvergente por SgcF ofrece potencial para la ingeniería de las hidrolasas de epóxido.
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