SummaryA wide variety of environmental contaminants exert estrogenic actions in wildlife, laboratory animals, and in human beings through binding to nuclear estrogen receptors (ERs). Here, the mechanism(s) of bisphenol A (BPA) to induce cell proliferation and the occurrence of its bioremediation by treatment with laccase are reported. BPA, highly present in natural world and considered as a model of environmental estrogen action complexity, promotes human cancer cell proliferation via ERa-dependent signal transduction pathways. Similar to 17b-estradiol, BPA increases the phosphorylation of both extracellular regulated kinase and AKT. Specific inhibitors of these kinase completely block the BPA effect on cancer cell proliferation. Notably, high BPA concentrations (i.e., 0.1 and 1 mM) are cytotoxic even in ERa-devoid cancer cells, indicating that an ERa-independent mechanism participates to BPA-induced cytotoxicity. On the other hand, BPA oxidation by laccase impairs the binding of this environmental estrogen to ERa loosing at all ERa-dependent effect on cancer cell proliferation. Moreover, the laccase-catalyzed oxidation of BPA reduces the BPA cytotoxic effect. Thus, laccase appears to impair BPA action(s), representing an invaluable bioremediation enzyme.2008 IUBMB IUBMB Life, 60(12): 843-852, 2008

The occurrence of tandem damage, due to reductive radical stress involving proteins and lipids, is shown by using a biomimetic model. It is made of unsaturated lipid vesicle suspensions in phosphate buffer in the presence of methionine, either as a single amino acid or as part of a protein such as RNase A, which contains four methionine residues. The radical process starts with the formation of H(.) atoms by reaction of solvated electrons with dihydrogen phosphate anions, which selectively attack the thioether function of methionine. The modification of methionine to alpha-aminobutyric acid is accompanied by the formation of thiyl radicals, which in turn cause the isomerization of the cis fatty acid residues to the trans isomers. The relationship between methionine modification and lipid damage and some details of the reductive radical stress obtained by proteomic analysis of irradiated RNase A are presented.

Lipidomics research, which focuses on the global changes in lipid metabolites, has recently been concerned with the type and roles of unsaturated lipids in the biological environment. The structural change induced by their conversion from the naturally occurring cis fatty acid geometry to the more thermodynamically stable trans configuration can affect membrane arrangement as well as lipid metabolism.[1] In the biomimetic model of thiyl radical-catalyzed isomerization of cis phospholipids, it was shown that when thiyl radicals are generated in the aqueous compartment and are able to diffuse in the lipid bilayer, then the interaction with unsaturated fatty acyl chains efficiently produces trans double bonds.[2] These findings suggested that radical-based degradation of sulfur-containing amino acid residues that are known to release diffusible thiol molecules could be the primer for tandem radical damage involving protein and lipid domains. We modeled such damage using g irradiation of lipid vesicle suspensions containing bovine pancreatic ribonuclease A (RNase A). The reaction of this protein with HC atoms was studied, and the inactivation was connected to the specific damage of sulfur moieties with release of low-molecular-weight thiols. [3] Liposomes were prepared by using dioleoyl phosphatidyl choline (DOPC) in the form of large unilamellar vesicles (LUVET) of 100 nm diameter.[4] The protein was added to the LUVET suspension and saturated with N 2 O prior to irradiation at a dose rate of 14.5 Gy min À1. 100 mL aliquots of the suspension were withdrawn at different irradiation times, over the interval of 2-70 min, for lipid isolation and derivatization to the corresponding fatty acid methyl esters.[5] This was followed by GC analysis to determine the cis/trans ratio. The solid circles in Figure 1 show the percentage of trans isomers (elaidate residues) formed as a function of irradiation dose. Control experiments in the absence of RNase A or by replacing RNase A with a protein without sulfur-containing amino acids, such as histone H1 type IIA from calf thymus, did not show any isomerization.In parallel, we followed changes in the enzyme activity, [6] as well as the transformation of the sulfur moieties by Raman spectroscopy, using lyophylized samples of aqueous solutions of native and irradiated RNase A. As expected from the radiation-induced inactivation, [3] a residual enzymatic activity of 67 % was found after exposure to only 33.3 Gy, followed by a slower decrease of the activity, which reached 50 % after 500 Gy. In the Raman spectra, the SÀS and CÀS stretching bands are visible in the 420-780 cm À1 spectral range. Native RNase A has four disulfide bridges that give rise to two different disulfide bands (n S-S ) at 514 and 535 cm À1

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