No abstract

The enzyme urate oxidase catalyzes the conversion of uric acid to 5-hydroxyisourate, one of the steps in the ureide pathway. Arthrobacter globiformis urate oxidase (AgUOX) was crystallized and structures of crystals soaked in the substrate uric acid, the inhibitor 8-azaxanthin and allantoin have been determined at 1.9-2.2 A resolution. The biological unit is a homotetramer and two homotetramers comprise the asymmetric crystallographic unit. Each subunit contains two T-fold domains of betabetaalphaalphabetabeta topology, which are usually found in purine- and pterin-binding enzymes. The uric acid substrate is bound tightly to the enzyme by interactions with Arg180, Leu222 and Gln223 from one subunit and with Thr67 and Asp68 of the neighbouring subunit in the tetramer. In the other crystal structures, lithium borate, 8-azaxanthin and allantoate are bound to the enzyme in a similar manner as uric acid. Based on these AgUOX structures, the enzymatic reaction mechanism of UOX has been proposed.

The growth rate and liver somatic index were significantly (P<0·05) lower in Mystus nemurus exposed to hydrogen sulphide compared to controls. These differences increased with corresponding increases in hydrogen sulphide concentrations. No significant differences (P>0·05) in Fulton's condition factor were detected between the exposed fish and the controls. The results revealed that liver somatic index is a more sensitive indicator of hydrogen sulphide toxicity compared to Fulton's condition factor.1998 The Fisheries Society of the British Isles

The crystal structures of pyruvate oxidase from Aerococcus viridans (AvPOX) complexed with flavin adenine dinucleotide (FAD), with FAD and thiamine diphosphate (ThDP) and with FAD and the 2-acetyl-ThDP intermediate (AcThDP) have been determined at 1.6, 1.8 and 1.9 A resolution, respectively. Each subunit of the homotetrameric AvPOX enzyme consists of three domains, as observed in other ThDP-dependent enzymes. FAD is bound within one subunit in the elongated conformation and with the flavin moiety being planar in the oxidized form, while ThDP is bound in a conserved V-conformation at the subunit-subunit interface. The structures reveal flexible regions in the active-site tunnel which may undergo conformational changes to allow the entrance of the substrates and the exit of the reaction products. Of particular interest is the role of Lys478, the side chain of which may be bent or extended depending on the stage of catalysis. The structures also provide insight into the routes for electron transfer to FAD and the involvement of active-site residues in the catalysis of pyruvate to its products.

D-3-Hydroxybutyrate dehydrogenase, which catalyzes the reversible reaction between D-3-hydroxybutyrate and acetoacetate, has been classified into the short-chain dehydrogenase/reductase family and is a useful marker in the assay of diabetes mellitus and/or ketoacidosis. The enzyme from Alcaligenes faecalis was crystallized in the apo form and in the holo form with acetate as a substrate analogue. The crystal structures of both forms were determined at 2.2 angstroms resolution. The enzyme is a tetramer composed of four subunits assembled with noncrystallographic 222 point symmetry. Each subunit has two domains. The principal domain adopts the Rossmann fold essential for nucleotide binding, which is a common feature of the SDR family. NAD+ is bound in a large cleft in the domain. The pyrophosphate group of NAD+ is covered by the small additional domain, which is supported by two extended arms allowing domain movement. In the catalytic site, a water molecule is trapped by the catalytic Tyr155 and Ser142 residues in the vicinity of the bound NAD+ and acetate. The substrate analogue acetate is bound above the nicotinamide plane. A substrate (D-3-hydroxybutylate) bound model can reasonably be constructed by adding two C atoms into the void space between the water O atom and the methyl group of the acetate, suggesting a substrate-bound state before enzymatic reaction occurs. Based on these structural features, a reaction mechanism has been proposed.