Chapter 3. Aims of the present study

The complete sequencing of the genome of baker’s yeast, S. cerevisiae, revealed that it has three genes encoding possible members of the hydratase/isomerase superfamily. The initial goal of the study was to start identifying the functions of these gene products and, especially, to search for the Δ32-enoyl-CoA isomerase or the Δ3,5–Δ2,4-dienoyl-CoA isomerase of S. cerevisiae. To start the search, the first aim was:

  1. To subclone the yeast gene YLR284c into a bacterial expression vector, to overexpress it and to determine the enzymatic activity of the gene product.

Once the gene YLR284c had been found to encode for a Δ32-enoyl-CoA isomerase, more specific aims for the study were set:

  1. To purify and characterize the recombinant yeast Δ32-enoyl-CoA isomerase.

  2. To determine the crystal structure of the Δ32-enoyl-CoA isomerase in the absence and presence of an active site ligand.

  3. To compare the Δ32-enoyl-CoA isomerase structures with each other and with the other known structures belonging to the hydratase/isomerase superfamily.