Structure of Cu2(indomethacin)4 and the reaction with superoxide in aprotic systems

The copper complex of indomethacin (1-(p-chlorobenzoyl)-5-methoxy-2-methyl-indole acetate), a common anti-inflammatory drug, was prepared and characterized. Crystal structure determination revealed the dimeric form of the 1:2 complex, namely Cu₂(indomethacin)₄ · L₂, in the unit cell. Suprisingly, the copper-copper distance (263 pm) was very close to metallic copper (256 pm). The two coordination sites in the copper-copper axis can be readily replaced by superoxide. An intriguing similarity to Cu₂(acetate)₄ was seen.Due to the lipophilic nature of the indomethacin ligand, this copper complex reacted with superoxide in aprotic solvents. The superoxide dismutating activity was successfully demonstrated in Me₂SO/water and acetonitrile/water mixtures using the nitro-blue tetrazolium assay and pulse radiolysis. The second-order rate constant of 6 · 10⁹ M^?1 · s^?1 in strictly aqueous systems dropped only slightly to 1.1 · 10⁹ M^?1 · s^?1 when aprotic solvents were used. This is the fastest rate constant ever observed for a copper-dependent dismutation of superoxide. The KO₂-induced lipid peroxidation in both erythrocytes and liver microsomes was suppressed by 70% in the presence of 1 · 10^?10 mol · ml^?1 of Cu₂(indomethacin)₄. The inhibitory action dropped to 25% when Cu₂Zn₂superoxide dismutase was employed. The formation of copper · indomethacin in rat serum after administration of indomethacin was shown in vitro and in vivo.     

Rabbit small intestine does not contain an annexin II/caveolin 1 complex as a target for 2-azetidinone cholesterol absorption inhibitors

Intestinal cholesterol absorption is specifically inhibited by the 2-azetidinone cholesterol absorption inhibitor ezetimibe. Photoreactive ezetimibe analogues specifically label a 145-kDa protein in the brush border membrane of enterocytes from rabbit small intestine identified as aminopeptidase N (CD13). In zebrafish and mouse small intestinal cytosol, a heterocomplex of M_r 52 kDa between annexin II and caveolin 1 was suggested as a target of ezetimibe. In contrast, in the cytosol and brush border membrane vesicles (BBMV) from rabbit small intestine of control animals or rabbits treated with the nonabsorbable cholesterol absorption inhibitor AVE 5530, both annexin II and caveolin 1 were exclusively present as monomers without any heterocomplex formation. Upon immunoprecipitation with annexin II a 52-kDa band was observed after immunostaining with annexin II antibodies, whereas no staining of a 52-kDa band occurred with anti-caveolin 1 antibodies. Vice versa, a 52-kDa band obtained by immunoprecipitation with caveolin 1 antibodies did not stain with annexin II-antibodies. The intensity of the 52-kDa band was dependent on the amount of antibody and was also observed with anti-actin or anti-APN antibodies suggesting that the 52-kDa band is a biochemical artefact. After incubation of cytosol or BBMV with radioactively labelled ezetimibe analogues, no significant amounts of the ezetimibe analogues could be detected in the immunoprecipitate with caveolin-1 or annexin II antibodies. Photoaffinity labelling of rabbit small intestinal BBMV with ezetimibe analogues did not result in labelling of proteins being immunoreactive with annexin II, caveolin 1 or a 52-kDa heterocomplex. These findings indicate that the rabbit small intestine does not contain an annexin II/caveolin 1 heterocomplex as a target for ezetimibe.     

Cell adhesion molecule in the hexactinellid Aphrocallistes vastus

Abstract. The Hexactinellida sponge Aphrocallistes vastus contains a soluble aggregation factor (AF) whose purification has been described in this communication. It is characterized by a S°_20.w value of 37 and a buoyant density of 1.45 g/cm³. The AF is a glycoporteinaceous particle composed of three major protein species; no core structure could be visualized. In the presence of Ca²⁺, the AF causes secondary aggregation of single cells. The aggregation process is temperature, pH, and ionic strength independent within a broad range. Evidence is presented indicating that two (or more) AF molecules are required for the establishment of a stable cell: cell interaction. In contrast to the AFs from demosponges, the hexactinellid AF functions species-unspecifically.     

A familial mutation in the testis-determining gene SRY shared by both sexes

A familial mutation in SRY, the gene coding for the testis-determining factor TDF, was identified in an XY female with gonadal dysgenesis, her father, her two brothers and her uncle. The mutation consists of a T to C transition in the region of the SRY gene coding for a protein motif known as the high mobility group (HMG) box, a protein domain known to confer DNA-binding specificity on the SRY protein. This point mutation results in the substitution, at amino acid position 109, of a serine residue for phenylalanine, a conserved aromatic residue in almost all HMG box motifs known. This F109S mutation was not found in 176 male controls. When recombinant wildtype SRY and SRY^F109S mutant protein were tested in vitro for binding to the target site AAC AAAG, no differences in DNA-binding activity were observed. These results imply that the F109S mutation either is a rare neutral sequence variant, or produces an SRY protein with slightly altered in vivo activity, the resulting sex phenotype depending on the genetic back-ground or environmental factors.This paper is dedicated by G. S. to Professor Ulrich Wolf on the occasion of his 60th birthday     

Die Anionenaustauschersubstanzen Aluminium- und Eisen (3)-hydroxyd-silikat-phosphat-humat Als Minerale Des Bodens

Zusammenfassung In wäßriger Lösung wurden Aluminium- oder Eisen(3)-Ionen bei Siedetemperatur mit Hydroxyl- und Silikat- oder Phosphat- oder Humat-Ionen unter ständiger Aufrechterhaltung bestimmter Mengenverhältnisse umgesetzt. In den entstandenen Niederschlägen wurden die Molarprozente an Hydroxyl-, Silikat-, Phosphat- und Humat-Ionen inweitem Bereich abhängig von den Konzentrationen dieser Ionen im Gleichgewicht gefunden. Daher sind diese Niederschläge als Anionenaustauscher anzusehen. Es wurde die Vermutung ausgesprochen, daß die Aluminium- und Eisen-Verbindungen des Bodens die gleichen Austauschersubstanzen darstellen, und versucht, diese Vermutung durch Besprechung einiger bodenkundlicher Befunde zu stützen. Es zeigte sich, daß der für die Pflanzenernährung wichtige, schnelle Anionenaustausch an diesen röntgenamorphen Austauschersubstanzen erfolgt, ebenso wie der Kationenaustausch über die in den 30er Jahren entdeckten, kristallinen Tonminerale verläuft.     

The lymphocyte restimulation system: Evaluation by intra-HLA-D group priming

Homozygous typing cells (HTC) were primed, using responding and stimulating lymphocytes of the same HLA-D groups. These intra-HLA-D group primings showed strong specific responses. Restimulation by HLA-D heterozygous and homozygous cell panels showed no correlation between the restimulating determinant and HLA-D. On the other hand, an unrelated individual, not carrying Dw4 and primed to Dw4 HTC, is restimulated by three of four Dw4-HTC. Thus, one non-HLA-D-associated restimulating determinant and another HLA-D-associated determinant could be identified. The differences among the four Dw4 HTC recognized in secondary MLC could reflect either recognition of separate gene products or recognition of separate determinants on the same gene product.