Title: First study on the determination of baseline biomarkers in Mimachlamys varia for an intra-port environmental biomonitoring in French Atlantic coastline (La Rochelle) Barbarin M, Muttin F, Thomas H Ref: Mar Pollut Bull, 182:113979, 2022 : PubMed
The characterization of organic and inorganic environmental pollution in coastal ecosystems, such as port areas, is complex and difficult to carry out due to the effect of environmental variables, as well as anthropic activities. For this study, the objective was to define a statistical method, taking into account the confounding factors influence, to define reference values for biomarkers in the black scallop (Mimachlamys varia). Thus, for five biomarkers (SOD (Superoxyde Dismutase), GST (Glutathion-S Transferase), MDA (Malondialdehyde), AChE (Acetylcholinesterase) and LAC (Laccase)), reference data could be described for individuals placed on sites more or less strongly impacted by specific environmental contaminations in port areas for more than two years, which had never been done before. All these results enabled us to calibrate and validate our approach in terms of active biomonitoring for the evaluation of a good ecological status of the environment of a port located on the French Atlantic coast.
In port areas the identification of contamination sources is necessary for an efficient management. Biomonitoring provides information on the environmental impact of the pollutants. It is often difficult to differentiate the natural variations of biomarkers from those induced by pollution. The present study aims to define a baseline level for biochemical biomarkers in limpet (Patella sp.) collected in four North-Corsica port areas. Reference data for five biomarkers (superoxide dismutase, glutathione S-transferase, laccase, pyruvate kinase and acetylcholinesterase) were described in a model, using length of the limpet shell, temperature and salinity. The measured biomarkers responses on potentially polluted sites usually fell within the range of the expected values for an unaffected site, suggesting that a main part of the variations is explained by environmental conditions. Not included in the model, biological factors (sex, development stage, etc.), annual variation or other physico-chemical parameter could explain the variations in the model.
Title: Isolation and characterization of a cDNA encoding rat liver cytosolic epoxide hydrolase and its functional expression in Escherichia coli Knehr M, Thomas H, Arand M, Gebel T, Zeller HD, Oesch F Ref: Journal of Biological Chemistry, 268:17623, 1993 : PubMed
A cDNA of 1992 base pairs encoding the complete rat liver cytosolic epoxide hydrolase has been isolated using a polymerase chain reaction-derived DNA fragment (Arand, M., Knehr, M., Thomas, H., Zeller, H. D., and Oesch, F. (1991) FEBS Lett. 294, 19-22) known to represent the 3'-end of the cytosolic epoxide hydrolase mRNA. Sequence analysis revealed an open reading frame of 1662 nucleotides corresponding to 554 amino acids (M(r) = 62,268). The DNA sequence obtained did not display significant homology to the sequences of microsomal epoxide hydrolase or leukotriene A4 hydrolase or to any other DNA included in the EMBL Data Bank (release 32). On Northern blotting of rat liver RNA, a single mRNA species was detected that was strongly induced on treatment of the animal with fenofibrate, a potent peroxisome proliferator. The most significant structure of the deduced protein is a modified peroxisomal targeting signal (Ser-Lys-Ile) at the carboxyl terminus that is regarded to be responsible for the unusual dual localization of the cytosolic epoxide hydrolase in peroxisomes as well as in the cytosol. In addition, a leucine zipper-like motif was identified at the amino terminus. Its possible implication for the observed dimeric structure of cytosolic epoxide hydrolase is discussed. The isolated cDNA was expressed in bacteria to yield a catalytically active enzyme. Specific activity of the crude lysate obtained exceeded that of rat liver cytosols from maximally induced animals by a factor of 8.
Title: An impaired peroxisomal targeting sequence leading to an unusual bicompartmental distribution of cytosolic epoxide hydrolase Arand M, Knehr M, Thomas H, Zeller HD, Oesch F Ref: FEBS Letters, 294:19, 1991 : PubMed
To gain an understanding of the mechanism by which the subcellular distribution of cytosolic epoxide hydrolase (cEH) is directed, we have analyzed the carboxy terminal region of rat liver cEH by means of cDNA cloning to define the structure of its possible peroxisomal targeting sequence (PTS). Purified cEH was subjected to peptide analysis following endoproteinase Glu-C digestion and HPLC-separation of the fragments. The obtained sequence information was used to perform PCR experiments resulting in the isolation of a 680 bp cDNA clone encoding the carboxy terminus of cEH. The deduced amino acid sequence displays a terminal tripeptide Ser-Lys-Ile which is highly homologous to the PTS (Ser-Lys-Leu) found in other peroxisomal enzymes. This slight difference appears to be sufficient to convert the signal sequence into an impaired and therefore ambivalent PTS, directing the enzyme partly to the peroxisomes and allowing part to reside in the cytosol.
Human liver epoxide hydrolases were characterized by several criteria and a cytosolic cis-stilbene oxide hydrolase (cEHCSO) was purified to apparent homogeneity. Styrene oxide and five phenylmethyloxiranes were tested as substrates for human liver epoxide hydrolases. With microsomes activity was highest with trans-2-methylstyrene oxide, followed by styrene 7,8-oxide, cis-2-methylstyrene oxide, cis-1,2-dimethylstyrene oxide, trans-1,2-dimethylstyrene oxide and 2,2-dimethylstyrene oxide. With cytosol the same order was obtained for the first three substrates, whereas activity with 2,2-dimethylstyrene oxide was higher than with cis-1,2-dimethylstyrene oxide and no hydrolysis occurred with trans-1,2-dimethylstyrene oxide. Generally, activities were lower with cytosol than with microsomes. The isoelectric point for both microsomal styrene 7,8-oxide and cis-stilbene oxide hydrolyzing activity was 7.0, whereas cEHCSO had an isoelectric point of 9.2 and cytosolic trans-stilbene oxide hydrolase (cEHTSO) of 5.7. The cytosolic epoxide hydrolases could be separated by anion-exchange chromatography and gel filtration. The latter technique revealed a higher molecular mass for cEHCSO than for cEHTSO. Both cytosolic epoxide hydrolases showed higher activities at pH 7.4 than at pH 9.0, whereas the opposite was true for microsomal epoxide hydrolase. The effects of ethanol, methanol, tetrahydrofuran, acetonitrile, acetone and dimethylsulfoxide on microsomal epoxide hydrolase depended on the substrate tested, whereas both cytosolic enzymes were not at all, or only slightly, affected by these solvents. Effects of different enzyme modulators on microsomal epoxide hydrolase also depended on the substrates used. Trichloropropene oxide and styrene 7,8-oxide strongly inhibited cEHCSO whereas cEHTSO was moderately affected by these compounds. Immunochemical investigations revealed a close relationship between cEHCSO and rat liver microsomal, but not cytosolic, epoxide hydrolase. Interestingly, cEHTSO has no immunological relationship to rat microsomal, nor to rat cytosolic epoxide hydrolase. cEHTSO from human liver differed also from its counterpart in the rat in that it was only moderately affected by tetrahydrofuran, acetonitrile and trichloropropene oxide. Five steps were necessary to purify cEHCSO. The enzyme has a molecular mass (49 kDa) identical to that of rat liver microsomal epoxide hydrolase.
