Structure of anabolic ornithine carbamoyltransferase from Campylobacter jejuni at 2.7 Å resolution.

Anabolic ornithine transcarbamoylase (aOTC) catalyzes the reaction between carbamoyl phosphate (CP) and L-ornithine (ORN) to form L-citrulline and phosphate in the urea cycle and L-arginine biosynthesis. The crystal structure of unliganded aOTC from Campylobacter jejuni (Cje aOTC) was determined at 2.7 Å resolution and refined to an R(work) of 20.3% and an R(free) of 24.0%. Cje aOTC is a trimer that forms a head-to-head pseudohexamer in the asymmetric unit. Each monomer is composed of an N-terminal CP-binding domain and a C-terminal ORN-binding domain joined by two interdomain helices. The Cje aOTC structure presents an open conformation of the enzyme with a relatively flexible orientation of the ORN-binding domain respective to the CP-binding domain. The conformation of the B2-H3 loop (residues 68-78), which is involved in binding CP in an adjacent subunit of the trimer, differs from that seen in homologous proteins with CP bound. The loop containing the ORN-binding motif (DxxxSMG, residues 223-230) has a conformation that is different from those observed in unliganded OTC structures from other species, but is similar to those in structures with bound ORN analogs. The major differences in tertiary structure between Cje aOTC and human aOTC are described.

ATP-dependent DNA ligase from Thermococcus sp. 1519 displays a new arrangement of the OB-fold domain.

DNA ligases join single-strand breaks in double-stranded DNA by catalyzing the formation of a phosphodiester bond between adjacent 5'-phosphate and 3'-hydroxyl termini. Their function is essential for maintaining genome integrity in the replication, recombination and repair of DNA. High flexibility is important for the function of DNA ligase molecules. Two types of overall conformations of archaeal DNA ligase that depend on the relative position of the OB-fold domain have previously been revealed: closed and open extended conformations. The structure of ATP-dependent DNA ligase from Thermococcus sp. 1519 (LigTh1519) in the crystalline state determined at a resolution of 3.02 Šshows a new relative arrangement of the OB-fold domain which is intermediate between the positions of this domain in the closed and the open extended conformations of previously determined archaeal DNA ligases. However, small-angle X-ray scattering (SAXS) measurements indicate that in solution the LigTh1519 molecule adopts either an open extended conformation or both an intermediate and an open extended conformation with the open extended conformation being dominant.

Investigation of formate transport through the substrate channel of formate dehydrogenase by steered molecular dynamics simulations.

Steered molecular dynamics simulation has revealed the mechanism of formate transport via the substrate channel of formate dehydrogenase. It is shown that the structural organization of the channel promotes the transport of formate anion in spite of the fact that the channel is too narrow even for such a small molecule. The conformational mobility of Arg284 residue, one of the residues forming the wall of the substrate channel, provides for the binding and delivery of formate to the active site.

Structures of the apo and holo forms of formate dehydrogenase from the bacterium Moraxella sp. C-1: towards understanding the mechanism of the closure of the interdomain cleft.

NAD(+)-dependent formate dehydrogenase (FDH) catalyzes the oxidation of formate ion to carbon dioxide coupled with the reduction of NAD(+) to NADH. The crystal structures of the apo and holo forms of FDH from the methylotrophic bacterium Moraxella sp. C-1 (MorFDH) are reported at 1.96 and 1.95 A resolution, respectively. MorFDH is similar to the previously studied FDH from the bacterium Pseudomonas sp. 101 in overall structure, cofactor-binding mode and active-site architecture, but differs in that the eight-residue-longer C-terminal fragment is visible in the electron-density maps of MorFDH. MorFDH also differs in the organization of the dimer interface. The holo MorFDH structure supports the earlier hypothesis that the catalytic residue His332 can form a hydrogen bond to both the substrate and the transition state. Apo MorFDH has a closed conformation of the interdomain cleft, which is unique for an apo form of an NAD(+)-dependent dehydrogenase. A comparison of the structures of bacterial FDH in open and closed conformations allows the differentiation of the conformational changes associated with cofactor binding and domain motion and provides insights into the mechanism of the closure of the interdomain cleft in FDH. The C-terminal residues 374-399 and the substrate (formate ion) or inhibitor (azide ion) binding are shown to play an essential role in the transition from the open to the closed conformation.

Atomic Resolution Crystal Structure of NAD(+)-Dependent Formate Dehydrogenase from Bacterium Moraxella sp. C-1.

The crystal structure of the ternary complex of NAD+-dependent formate dehydrogenase from the methylotrophic bacterium Moraxella sp. C-1 with the cofactor (NAD+) and the inhibitor (azide ion) was established at 1.1 A resolution. The complex mimics the structure of the transition state of the enzymatic reaction. The structure was refined with anisotropic displacitalicents parameters for non-hydrogen atoms to a R factor of 13.4%. Most of the nitrogen, oxygen, and carbon atoms were distinguished based on the analysis of the titalicperature factors and electron density peaks, with the result that side-chain rotamers of histidine residues and most of asparagine and glutamine residues were unambiguously determined. A comparative analysis of the structure of the ternary complex determined at the atomic resolution and the structure of this complex at 1.95 A resolution was performed. In the atomic resolution structure, the covalent bonds in the nicotinamide group are somewhat changed in agreitalicent with the results of quantum mechanical calculations, providing evidence that the cofactor acquires a bipolar form in the transition state of the enzymatic reaction.