PDBsum entry 1ipe

Oxidoreductase

PDB id: 1ipe

Contents

Protein chains

259 a.a. *

Ligands

NDP ×2

Waters ×61

* Residue conservation analysis

References listed in PDB file

Key reference

Title

Capturing enzyme structure prior to reaction initiation: tropinone reductase-Ii-Substrate complexes.

Authors

A.Yamashita, M.Endo, T.Higashi, T.Nakatsu, Y.Yamada, J.Oda, H.Kato.

Ref.

Biochemistry, 2003, 42, 5566-5573. [DOI no: 10.1021/bi0272712]

PubMed id

12741812

Abstract

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.

Secondary reference #1

Title

Structure of tropinone reductase-Ii complexed with NADP+ and pseudotropine at 1.9 a resolution: implication for stereospecific substrate binding and catalysis.

Authors

A.Yamashita, H.Kato, S.Wakatsuki, T.Tomizaki, T.Nakatsu, K.Nakajima, T.Hashimoto, Y.Yamada, J.Oda.

Ref.

Biochemistry, 1999, 38, 7630-7637. [DOI no: 10.1021/bi9825044]

PubMed id

10387002

Secondary reference #2

Title

Crystal structures of two tropinone reductases: different reaction stereospecificities in the same protein fold.

Authors

K.Nakajima, A.Yamashita, H.Akama, T.Nakatsu, H.Kato, T.Hashimoto, J.Oda, Y.Yamada.

Ref.

Proc Natl Acad Sci U S A, 1998, 95, 4876-4881. [DOI no: 10.1073/pnas.95.9.4876]

PubMed id

9560196

Figure 2.
Fig. 2. (A) Structures of TR dimers. Subunits of TR-I (green and blue) and TR-II (yellow and red) are related, respectively, by a noncrystallographic and crystallographic twofold axis positioned at the center of each dimer and oriented perpendicular to the plane of the figure. NADP+ bound in the TR-I subunits is shown by ball-and-stick models. Disordered regions in chain A of TR-I and both of the TR-II subunits are shown by dots. (B) C traces of TR-I (green) and TR-II (orange) subunits are superimposed by the program LSQKAB (9) using all possible C pairs. The binding position of NADP+ in TR-I and the side chains of the three catalytic residues also are shown. The small lobes are shown in a gray background. Arrows indicate the clefts formed between the core domain and the small lobe (A and B). The figures were prepared by using the programs MOLSCRIPT (13) and RASTER3D (14).

Figure 5.
Fig. 5. Environments of the substrate binding sites and the binding models of tropinone. (A and B) Inner molecular surfaces of the tropinone binding pockets of TR-I (A) and TR-II (B). Electrostatic charge distributions are shown on the molecular surfaces in blue (positive charge) and red (negative charge). Note that the inner surface of TR-II (B) is somewhat deformed at the lower left corner due to the absence of several residues that constitute the small lobe. (C and D) Amino acid side chains that form the tropinone binding pockets viewed from the same direction as in A and B. Two TR-II residues (D) corresponding to the Leu-208 and Val-209 of TR-I (C) are missing in this structural model. Bonds from the carbon atoms in tropinone are shown in yellow, whereas the protein bonds from the C atoms are green to half their lengths. This figure was prepared by the GRASP program (23).