No abstract

No abstract

Background During the last decades, a worldwide increase in the number of cases of depression accompanied by rising prescription rates of antidepressants was recorded. In Germany, the two most prescribed antidepressants are the selective serotonin reuptake inhibitor (SSRI) citalopram and the serotonin and noradrenalin reuptake inhibitor (SNRI) venlafaxine, taking about 30% of the market share. Both antidepressants are found frequently in surface waters and have the potential of adversely affecting aquatic organisms. Most studies dealing with antidepressants address apical endpoints and behaviour; however, only few studies investigate biochemical biomarkers and histopathological alterations. We conducted citalopram and venlafaxine exposure experiments over 5 months, starting with brown trout eggs in the eyed-ova stage, as well as with juvenile brown trout for 4 weeks. Exposure concentrations ranged from environmentally relevant 1 µg/L up to 1 mg/L. In this study, we investigated the effects of the antidepressants on b-esterase activity (neurotoxicity), stress protein level (proteotoxicity) and superoxide dismutase activity (oxidative stress). Additionally, we assessed the health status of the liver by means of histopathological analyses. Results We were able to show that both antidepressants did neither induce proteotoxic nor neurotoxic effects in brown trout. But for venlafaxine, the biochemical biomarker for oxidative stress (superoxide dismutase activity) was significantly increased in larvae exposed to at least 10-µg/L venlafaxine at 7 °C. With regard to liver histopathology, fish exposed to higher citalopram concentrations were in a worse health condition than control fish irrespective of their life stage. Also, the energy storage of fish exposed to 1 mg/L citalopram was reduced. Conclusion Thus, we here report citalopram-dependent histopathological alterations in brown trout liver, and the induction of oxidative stress by venlafaxine.

In terrestrial snails, thermal selection acts on shell coloration. However, the biological relevance of small differences in the intensity of shell pigmentation and the associated thermodynamic, physiological, and evolutionary consequences for snail diversity within the course of environmental warming are still insufficiently understood. To relate temperature‐driven internal heating, protein and membrane integrity impairment, escape behavior, place of residence selection, water loss, and mortality, we used experimentally warmed open‐top chambers and field observations with a total of >11,000 naturally or experimentally colored individuals of the highly polymorphic species Theba pisana (O.F. MÜLLER, 1774). We show that solar radiation in their natural Mediterranean habitat in Southern France poses intensifying thermal stress on increasingly pigmented snails that cannot be compensated for by behavioral responses. Individuals of all morphs acted neither jointly nor actively competed in climbing behavior, but acted similarly regardless of neighbor pigmentation intensity. Consequently, dark morphs progressively suffered from high internal temperatures, oxidative stress, and a breakdown of the chaperone system. Concomitant with increasing water loss, mortality increased with more intense pigmentation under simulated global warming conditions. In parallel with an increase in mean ambient temperature of 1.34°C over the past 30 years, the mortality rate of pigmented individuals in the field is, currently, about 50% higher than that of white morphs. A further increase of 1.12°C, as experimentally simulated in our study, would elevate this rate by another 26%. For 34 T. pisana populations from locations that are up to 2.7°C warmer than our experimental site, we show that both the frequency of pigmented morphs and overall pigmentation intensity decrease with an increase in average summer temperatures. We therefore predict a continuing strong decline in the frequency of pigmented morphs and a decrease in overall pigmentation intensity with ongoing global change in areas with strong solar radiation.

This paper describes the enzymatic synthesis of the C-terminal fragment H-Gly-Trp-Met-Asp-Phe-NH2 of cholecystokinin. Immobilized enzymes were used for the formation of all peptide bonds except thermolysin. Beginning the synthesis with phenylacetyl (PhAc) glycine carboxamidomethyl ester (OCam) and H-Trp-OMe by using immobilized papain as biocatalyst in buffered ethyl acetate, the dipeptide methyl ester was then coupled directly with Met-OEt.HCl by alpha-chymotrypsin/Celite 545 in a solvent free system. For the 3+2 coupling PhAc-Gly-Trp-Met-OEt had to be converted into its OCam ester. The other fragment H-Asp(OMe)-Phe-NH2 resulted from the coupling of Cbo-Asp(OMe)-OH with H-Phe-NH2.HCl and thermolysin as catalyst, followed by catalytic hydrogenation. Finally PhAc-Gly-Trp-Met-Asp-Phe-NH2 was obtained in a smooth reaction from PhAc-Gly-Trp-Met-OCam and H-Asp(OMe)-Phe-NH2 with alpha-chymotrypsin/Celite 545 in acetonitrile, followed by basic hydrolysis of the beta-methyl ester. The PhAc-group is removed with penicillin G amidase and CCK-5 is obtained in an overall isolated yield of 19.6%.

Polystyrene-polyoxyethylene craft copolymers have been used for step-wise peptide synthesis. After completion of synthesis the protecting groups are cleaved under acidic conditions, where the polymer-peptide bond is stable. These gels in comparison to polystyrene peptide gels, show better properties for applications in affinity chromatography as well as synthesis on solid supports, because the advantageous properties of polystyrene beads are combined with the excellent spacer behavior of polyoxyethylene chains (mobility, solvation by water and organic solvents). Peptide gels with polylysine sequences have been synthesized as highly selective stationary phases for the separation of the homologous oligo desoxyribonucleotides (dT)n with n = 1-3. The principal possibilities of these gels for affinity chromatography is demonstrated.

The preparation of apoferredoxin from spinach ferredoxin and ferredoxin of Clostridium pasteurianum with a,a'-dipyridyl without the addition of reducing agents is described.Reconstitution to form active ferredoxins can be achieved in high yields under various conditions. 2-Mercaptoethanol can be replaced by cysteamine or Cleland'sReagent. The reconstitution is also possible by using elemental sulfur, cysteamine, and iron(I1) ammonium sulfate. The origin of labile sulfur in Clostridial ferredoxin has been investigated. The elemental analyses show conclusively that in ferredoxin there are 5-6 additional sulfurs besides the 8 sulfurs from the cysteine moieties, while apoferredoxin contains only the 8 sulfurs of the cysteine groups. A clostridial [35S] ferredoxin has been prepared by using Na,a5S in the resynthesis. The uptake of sulfide corresponded to 6 equivalents of sulfur per mole of ferredoxin. Treatment of the %-labelled clostridial ferredoxin yielded apoferredoxin essentially free of radioactivity. From all of our analytical data the most probable formulation for Clostridium pasteurianum includes 8 equivalents of cysteine, 6 equivalents of labile sulfur, and 6 equivalents of iron.The hydropersulfides of cysteamine and cysteine methylester were prepared in solution. Paramagnetic complexes with g,,-values below 2 were prepared from the ferrous complex of cysteamine or cysteine methyl ester and hydropersulfides of cysteamine or cysteinc methyl ester. The g-values of the persulfide complexes are 1.92, 1.94 and 2.00. The monomeric unit of these paramagnetic compounds is formulated as a disulfide-iron(I1)-sulfide complex which is equivalent to a persulfide-thiolato-iron(II), and a sulfur complex of bis-thiolato-iron(I1).If one Considers the active center in plant ferredoxin as an analogous dimeric unit and the center in clostridial ferredoxin as an oligomeric unit, the labile sulfur may be represented in the form of S," or disulfide anion S,a-in oxidized ferredoxins or supersulfide anion S,*-in reduced ferredoxins. Structures are postulated for the active center of binuclear plant ferredoxins that are in agreement with the chemical and physical properties. These contain Fe-S,-Fe groups which are believed to be non-planar and exhibit stereochemistry similar to that found earlier by X-ray analysis for the binuclear complex (CO),FeS,Fe( CO),. The oxidized binuclear ferredoxins are described by the valence resonance hybrids Fe(II)SZ0Fe(II) and Fe(III)S,a-Fe(III), and the reduced paramagnetic form by Fe(1I)S;-Fe(I1) and Fe(II)Sz2-Fe(I1I). This seems to unite different theories concerning the cause for the characteristic ESR signal of ferredoxins.For polynuclear ferredoxins of the clostridial ferredoxin type an active center is suggested which contains units of binuclear ferredoxins in modified form. It appears that in the polynuclear active centers the formulation Fe(III)S,2-Fe(III) participates more in the ground state. I n the light of the chemical and physical properties it is proposed that in clostridial ferredoxin...

The restriction endonuclease EcoRI binds and cleaves DNA containing GAATTC sequences with high specificity. According to the crystal structure, most of the specific contacts of the enzyme to the DNA are formed by the extended chain region and the first turn of alpha-helix alpha 4 (amino acids 137-145). Here, we demonstrate that a dodecapeptide (WDGMAAGNAIER), which is identical in the underlined parts of its sequence to EcoRI amino acids 137-145, specifically binds to GAATTC sequences. The peptide inhibits DNA cleavage by EcoRI but not by BamHI, BclI, EcoRV, HindIII, PacI, and XbaI. DNA cleavage by XbaI is slowed down at sites that partially overlap with EcoRI sites. The peptide inhibits cleavage of GAATTC sites by ApoI, which recognizes the sequence RAATTY. It interferes with DNA methylation by the EcoRI methyltransferase but not by the BamHI methyltransferase. It competes with EcoRI for DNA binding. Based on these results, the DNA binding constant of the peptide to GAATTC sequences was calculated to be 3 x 10(4) M-1. DNA binding is not temperature-dependent, suggesting that binding of the peptide is entropy-driven. As the peptide does not show any nonspecific binding to DNA, its DNA binding specificity is similar to that of EcoRI, in spite of the fact that the affinity is much smaller. These results suggest that contacts to the phosphate groups in EcoRI mainly provide binding affinity, whereas the specificity of EcoRI is based to a large extent on sequence-specific base contacts.