W0038: Crystal Structure of Human Ornithine Transcarbamoylase Complexed with Carbamoyl Phosphate and L-Norvaline at 1.9 A Resolution

Crystal Structure of Human Ornithine Transcarbamoylase Complexed with Carbamoyl Phosphate and L-norvaline at 1.9 Å Resolution. Dashuang Shi ^†, Hiroki Morizono ^‡, Mika Aoyagi ^†, Mendel Tuchman ^‡ and Norma M. Allewell ^†*, ^†Department of Biochemistry, University of Minnesota, St. Paul, MN55108, USA. ^‡Department of Pediatrics, Medical School, University of Minnesota, Minneapolis, MN55455, USA.

The crystal structure of human ornithine transcarbamoylase (OTCase) complexed with carbamoyl phosphate (CP) and L-norvaline (NOR) has been determined to 1.9-Å resolution. Binding of CP produces some new interactions between CP and the enzyme although binding of the phosphate group of CP is similar to that of N-(phosphonoacetyl)-L-ornithine (PALO) [1]. 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 and side-chain carbonyl oxygen of Gln-171 and the side-chain of Arg-330. The carbonyl plane of CP rotates about 60( compared to the equivalent plane of PALO in PALO complexed OTCase. As a result, the side chain of NOR points vertically to this plane to optimize the attack to the carbonyl carbon of CP. The residues which interacts with NOR are similar to the residues which interact with the ornithine (ORN) moiety of PALO. The side-chain of NOR is well defined with the side-chains of Leu-163, Leu-200, Met-268 and Pro-305 forming a hydrophobic wall and also close to the side-chain of Cys-303. C-( atom of NOR is close to the carbonyl oxygen of Leu-304 (3.53 Å), S-( atom of Cys-303 (4.06 Å) and carbonyl carbon of CP (3.29 Å). Even though the N-( atom of ornithine is absent in this structure, the side-chain of NOR is positioned so as to enable the N-( atom of ornithine to be modeled in a position to donate a hydrogen to the S-( 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 also similar to that of the PALO-enzyme complex. The structure of the active sites in the complexes of aspartate transcarbamoylase (ATCase) with various substrates or inhibitors is similar to this OTCase structure, implying a common revolutionary origin.