Molecular architecture of the ankyrin SOCS box family of Cul5-dependent E3 ubiquitin ligases.

Multi-subunit Cullin-RING E3 ligases often use repeat domain proteins as substrate-specific adaptors. Structures of these macromolecular assemblies are determined for the F-box-containing leucine-rich repeat and WD40 repeat families, but not for the suppressor of cytokine signaling (SOCS)-box-containing ankyrin repeat proteins (ASB1-18), which assemble with Elongins B and C and Cul5. We determined the crystal structures of the ternary complex of ASB9-Elongin B/C as well as the interacting N-terminal domain of Cul5 and used structural comparisons to establish a model for the complete Cul5-based E3 ligase.

High-affinity inhibitors of human NAD-dependent 15-hydroxyprostaglandin dehydrogenase: mechanisms of inhibition and structure-activity relationships.

Background: 15-Hydroxyprostaglandin dehydrogenase (15-PGDH, EC 1.1.1.

The crystal structure of human GLRX5: iron-sulfur cluster co-ordination, tetrameric assembly and monomer activity.

Human GLRX5 (glutaredoxin 5) is an evolutionarily conserved thiol-disulfide oxidoreductase that has a direct role in the maintenance of normal cytosolic and mitochondrial iron homoeostasis, and its expression affects haem biosynthesis and erythropoiesis. We have crystallized the human GLRX5 bound to two [2Fe-2S] clusters and four GSH molecules. The crystal structure revealed a tetrameric organization with the [2Fe-2S] clusters buried in the interior and shielded from the solvent by the conserved β1-α2 loop, Phe⁶⁹ and the GSH molecules.

Kinase domain insertions define distinct roles of CLK kinases in SR protein phosphorylation.

Splicing requires reversible phosphorylation of serine/arginine-rich (SR) proteins, which direct splice site selection in eukaryotic mRNA. These phosphorylation events are dependent on SR protein (SRPK) and cdc2-like kinase (CLK) families. SRPK1 phosphorylation of splicing factors is restricted by a specific docking interaction whereas CLK activity is less constrained.

Structure-activity relationships of human AKR-type oxidoreductases involved in bile acid synthesis: AKR1D1 and AKR1C4.

Two members of the human aldo-keto reductase (AKR) superfamily participate in the biosynthesis of bile acids by catalyzing the NADP(H) dependent reduction of 3-keto groups (AKR1C4) and Delta4 double bonds (AKR1D1) of oxysterol precursors. Structure determination of human AKR1C4 and homology modelling of AKR1D1 followed by docking experiments were used to explore active site geometries. Substrate docking resulted in ligand poses satisfying catalytic constraints, and indicates a critical role for Trp227/230 in positioning the substrate in a catalytically competent orientation.

Ligand supplementation as a method to increase soluble heterologous protein production.

Ligand interactions are central to enzyme or receptor function, constituting a cornerstone in biochemistry and pharmacology. Here we discuss a ligand application that can be exploited to significantly increase the proportion of recombinant protein expressed in soluble form, by including ligands during the culture. Provided that a sufficiently soluble, cell-permeable and avid ligand is available, one can use it to stabilize nascently synthesized proteins, and in this manner promote solubility and prevent aggregation.

The molecular mechanism of nitrogen-containing bisphosphonates as antiosteoporosis drugs.

Osteoporosis and low bone mass are currently estimated to be a major public health risk affecting >50% of the female population over the age of 50. Because of their bone-selective pharmacokinetics, nitrogen-containing bisphosphonates (N-BPs), currently used as clinical inhibitors of bone-resorption diseases, target osteoclast farnesyl pyrophosphate synthase (FPPS) and inhibit protein prenylation. FPPS, a key branchpoint of the mevalonate pathway, catalyzes the successive condensation of isopentenyl pyrophosphate with dimethylallyl pyrophosphate and geranyl pyrophosphate.

Characterization of human DHRS6, an orphan short chain dehydrogenase/reductase enzyme: a novel, cytosolic type 2 R-beta-hydroxybutyrate dehydrogenase.

Human DHRS6 is a previously uncharacterized member of the short chain dehydrogenases/reductase family and displays significant homologies to bacterial hydroxybutyrate dehydrogenases. Substrate screening reveals sole NAD(+)-dependent conversion of (R)-hydroxybutyrate to acetoacetate with K(m) values of about 10 mm, consistent with plasma levels of circulating ketone bodies in situations of starvation or ketoacidosis. The structure of human DHRS6 was determined at a resolution of 1.

DNA-PKcs-dependent signaling of DNA damage in Dictyostelium discoideum.

DNA double-strand breaks (DSBs) can be repaired by either homologous recombination (HR) or nonhomologous end-joining (NHEJ). In vertebrates, the first step in NHEJ is recruitment of the DNA-dependent protein kinase (DNA-PK) to DNA termini. DNA-PK consists of a catalytic subunit (DNA-PKcs) that is recruited to DNA ends by the Ku70/Ku80 heterodimer.