Crystal structure of human ornithine transcarbamylase complexed with carbamoyl phosphate and L‐norvaline at 1.9 Å resolution

Abstract

The crystal structure of human ornithine transcarbamylase (OTCase) complexed with carbamoyl phosphate (CP) and L-norvaline (NOR) has been determined to 1.9-Å resolution. There are significant differences in the interactions of CP with the protein, compared with the interactions of the CP moiety of the bisubstrate analogue N-(phosphonoacetyl)-L-ornithine (PALO). The carbonyl plane of CP rotates about 60° compared with the equivalent plane in PALO complexed with OTCase. This positions the side chain of NOR optimally to interact with the carbonyl carbon of CP. The mixed-anhydride oxygen of CP, which is analogous to the methylene group in PALO, interacts with the guanidinium group of Arg-92; the primary carbamoyl nitrogen interacts with the main-chain carbonyl oxygens of Cys-303 and Leu-304, the side chain carbonyl oxygen of Gln-171, and the side chain of Arg-330. The residues that interact with NOR are similar to the residues that interact with the ornithine (ORN) moiety of PALO. The side chain of NOR is well defined and close to the side chain of Cys-303 with the side chains of Leu-163, Leu-200, Met-268, and Pro-305 forming a hydrophobic wall. C-δ of NOR is close to the carbonyl oxygen of Leu-304 (3.56 Å), S-γ atom of Cys-303 (4.19 Å), and carbonyl carbon of CP (3.28 Å). Even though the N-ϵ atom of ornithine is absent in this structure, the side chain of NOR is positioned to enable the N-ϵ of ornithine to donate a hydrogen to the S-γ atom of Cys-303 along the reaction pathway. Binding of CP and NOR promotes domain closure to the same degree as PALO, and the active site structure of CP-NOR-enzyme complex is similar to that of the PALO-enzyme complex. The structures of the active sites in the complexes of aspartate transcarbamylase (ATCase) with various substrates or inhibitors are similar to this OTCase structure, consistent with their common evolutionary origin. Proteins 2000;39:271–277. © 2000 Wiley-Liss, Inc.