| Δ3-Δ2-Enoyl-CoA isomerase from the yeast Saccharomyces cerevisiae: Molecular and structural characterization | ||
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The MAD datasets collected at the peak and remote wavelengths were scaled to the dataset collected at the inflection point wavelength with SCALEIT (Collaborative Computational Project Number 4 1994). The initial heavy-atom positions were located with RSPS (Collaborative Computational Project Number 4 1994) and SOLVE (Terwilliger & Berendzen 1999). The two heavy-atom positions found were refined and the phases to 2.5 Å resolution were calculated with the program MLPHARE (Collaborative Computational Project Number 4 1994). The initial maps were improved by solvent-flattening calculations and the phases were extended to 2.15 Å resolution with the DM program (Cowtan & Main 1996), after which the maps were easily interpretable with the O program (Jones et al. 1991). The model of Eci1p monomer was built manually in O. Subsequently, the model was subjected to restrained refinement, using the maximum-likelihood target, with REFMAC (Murshudov et al. 1997). Water molecules were added automatically by the solvent-building mode of the program ARP/wARP (Perrakis et al. 1999). Ethylene glycol and perrhenate molecules were also added into the refinement. The pdb entry code for the perrhenate-complexed yeast enoyl-CoA isomerase is 1HNU.
The unliganded structure (not soaked in perrhenate) of the yeast enoyl-CoA isomerase was solved by rigid body refinement with REFMAC using the perrhenate-complexed structure as the starting model. Further restrained refinement to 2.5 Å resolution was also carried out with REFMAC. The pdb entry code is 1HNO.