Dept. of Biochemistry & Organic Chemistry

Physiological functions of glutathione transferases:

role in steroidogenesis

• Human GSTA3-3 and biosynthesis of steroid hormones.
  Human GST A3-3 (hGST A3-3 is expressed selectively in steroidogenic organs and shows high steroid isomerase activity – a key chemical reaction in the biosynthesis of steroid hormones - towards precursors of progesterone and testosterone.
  RNA interference is used to knock down hGST A3-3 expression in cultured human cells. The ability of the transfected cells to produce steroid hormones is evaluated by measuring the amount of progesterone or testosterone released in culture medium.
• Identification of non-human mammalian alpha-class glutathione transferases involved in steroid hormone biosynthesis.
  GSTs with high sequence similarity to human hGST A3-3 are cloned, heterologously expressed and purified. Their enzymatic properties towards steroid and non-steroid substrates are analyzed. The animals considered are cow, pig, mouse and rat.
• Modulation of steroidogenesis through inhibition of alpha-class glutathione transferases.
  Potential inhibitors of GST–dependent steroid isomerase activity are identified and tested for their ability to affect steroidogenesis in cultured cells.

Methods:

Mammalian cell culture, gene knockdown, siRNA transfection, cell proliferation/toxicity assay, RT-PCR, western-blot, cloning, protein purification, enzymology.