Kinetic studies on the formation of sulfonyl radicals and their addition to carbon-carbon multiple bonds.

The reactions of α-hydroxyl and α-alkoxyl alkyl radicals with methanesulfonyl chloride (MeSO(2)Cl) have been studied by pulse radiolysis at room temperature. The alkyl radicals were produced by ionizing radiation of N(2)O-saturated aqueous solution containing methanol, ethanol, isopropanol, or tetrahydrofuran. The transient optical absorption spectrum consisted of a broad band in the region 280-380 nm with a maximum at 320 nm typical of the MeSO(2)(•) radical. The rate constants in the interval of 1.7 × 10(7)-2.2 × 10(8) M(-1) s(-1) were assigned to an electron-transfer process that leads to MeSO(2)Cl(•-), subsequently decaying into MeSO(2)(•) radical and Cl(-). The rate constants for the addition of CH(3)SO(2)(•) to acrolein and propiolic acid were found to be 4.9 × 10(9) M(-1) s(-1) and 5.9 × 10(7) M(-1) s(-1), respectively, in aqueous solutions and reversible. The reactivity of tosyl radical (p-CH(3)C(6)H(4)SO(2)(•)) toward a series of alkenes bearing various functional groups was also determined by competition kinetics in benzene. The rate constants for the addition of tosyl radical to alkenes vary in a much narrower range than the rate constants for the reverse reaction. The stabilization of the adduct radical substantially contributes to the increase of the rate constant for the addition of tosyl radical to alkenes and, conversely, retards the ß-elimination of tosyl radical.

Radical damage involving sulfur-containing enzymes and membrane lipids.

Free radicals induce protein modifications, often associated to many biological phenomena. This mini-review overviews the approaches we have used to elucidate the radical-induced damages on sulfur-containing enzymes, such as ribonuclease and lysozyme, and the possible post-translational mechanism of the damage to another cell compartment, such as lipid domain.

One-electron reduction of methanesulfonyl chloride. The fate of MeSO2Cl(.)(-) and MeSO2(.) intermediates in oxygenated solutions and their role in the cis-trans isomerization of mono-unsaturated fatty acids.

The one-electron reduction of methanesulfonyl chloride (MeSO2Cl) leads, in the first instance, to an electron adduct MeSO2Cl(.)(-) which lives long enough for direct detection and decays into sulfonyl radicals MeSO2(.) and Cl(-), with k = 1.5 x 10(6) s(-1). Both, MeSO2Cl(.)(-) and MeSO2(.) showed a similar absorption in the UV with lambdamax of 320 nm. In the presence of oxygen, MeSO2Cl(.)(-) transfers an electron to O(2) and establishes an equilibrium with superoxide. The rate constant for the forward reaction was measured to 4.1 x 10(9) M(-1) s(-1), while for the back reaction only an interval of 1.7 x 10(5) to 1.7 x 10(6) M(-1) s(-1) could be estimated, with a somewhat higher degree of confidence for the lower value. This corresponds to an equilibrium constant in the range of 2.4 x 10(3) to 2.4 x 10(4). With reference to E degrees (O2/O2(.)(-)) = -155 mV, the redox potential of the sulfonyl chloride couple, E degrees (MeSO2Cl/MeSO2Cl(.)(-)), thus results between being equal to -355 and -414 mV (vs NHE). MeSO2Cl(.)(-) reduces (besides O2) 4-nitroacetophenone. The underlying electron transfer took place with k = 1.5 x 10(9) M(-1) s(-1), corroborating an E degrees for the sulfonyl chloride couple significantly exceeding the above listed lower value. The MeSO2(.) radical added to oxygen with a rate constant of 1.1 x 10(9) M(-1) s(-1). Re-dissociation of O2 from MeSO2OO(.) occurred only very slowly, if at all, that is, with k << 10(5) s(-1). MeSO2(.) radicals can act as the catalyst for the cis-trans isomerization of several Z- and E-mono-unsaturated fatty acid methyl esters in homogeneous solution. The effectiveness of the isomerization processes has been addressed, and in the presence of oxygen the isomerization is completely suppressed.

The reaction of hydrogen atoms with methionine residues: A model of reductive radical stress causing tandem protein-lipid damage.

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.

Investigation of radical-based damage of RNase A in aqueous solution and lipid vesicles.

The gamma-irradiation of bovine pancreatic ribonuclease A (RNase A) in aqueous solution were investigated at different doses by vibrational spectroscopy as well as enzymatic assay, electrophoresis, and HPLC analysis. Both functional and structural changes of the protein were caused by attack of H(*) atoms and (*)OH radicals. In particular, Raman spectroscopy was shown to be a useful tool in identifying conformational changes of the protein structure and amino acidic residues that are preferential sites of the radical attack (i.e., tyrosine and methionine). After partial structural changes by the initial radical attack, the internal sulfur-containing amino acid residues were rendered susceptible to transformation. By using the biomimetic model of dioleoyl phosphatidyl choline vesicle suspensions containing RNase A, the damage to methione residues could be connected to a parallel alteration of membrane unsaturated lipids. In fact, thiyl radical species formed from protein degradation can diffuse into the lipid bilayer and cause isomerization of the naturally occurring cis double bonds. As a consequence, trans unsaturated fatty acids are formed in vesicles and can be considered to be markers of this protein damage.

Vibrational characterisation and biological activity of carnosine and its metal complexes.

The spectroscopic characterisation of free carnosine and its coordination compounds with Cu(II), Zn(II) and Co(II) ions are discussed. Raman and IR studies on metal-carnosine systems have been performed, obtaining a relationship between the vibrational spectra and the structure of the predominant species formed. The biological activity of free carnosine and of its complexes is briefly considered.