To understand the catalytic mechanism of an enzyme, it is crucial to determine the crystallographic structures corresponding to the individual reaction steps. Here, we report two crystal structures of enzyme - substrate complexes prior to reaction initiation: tropinone reductase-II (TR-II) NADPH and TR-II - NADPH - tropinone complexes, determined from the identical crystals. A combination of two kinetic crystallographic techniques, a continuous flow of the substrates and Laue diffraction measurements, enabled us to capture the transit structures prior to the reaction proceeding. A structure comparison of the enzyme - substrate complex elucidated in this study with the enzyme-product complex in our previous study indicates that one of the substrates, tropinone, is rotated relative to the product so as to make the spatial organization in the active site favorable for the reaction to proceed. Side chains of the residues in the active site also alter their conformations to keep the complementarity of the space for the substrate or the product and to assist the rotational movement.
|Number of pages||8|
|Publication status||Published - May 20 2003|
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