Sulfoxidation mechanism of vanadium bromoperoxidase from Ascophyllum nodosum. Evidence for direct oxygen transfer catalysis.

We have previously shown that vanadium bromoperoxidase from Ascophyllum nodosum mediates production of the (R)-enantiomer of methyl phenyl sulfoxide with 91% enantiomeric excess. Investigation of the intrinsic selectivity of vanadium bromoperoxidase reveals that the enzyme catalyzes the sulfoxidation of methyl phenyl sulfide in a purely enantioselective manner. The K(m) of the enzyme for methyl phenyl sulfide was determined to be approximately 3.5 mM in the presence of 25% methanol or tert-butanol. The selectivity of the sulfoxidation of methyl phenyl sulfide is optimal in the temperature range 25-30 degrees C and can be further optimized by increasing the enzyme concentration, yielding selectivities with up to 96% enantiomeric excess. Furthermore, we established for the first time that vanadium bromoperoxidase is functional at temperatures up to 70 degrees C. A detailed investigation of the sulfoxidation activity of this enzyme using (18)O-labeled hydrogen peroxide shows that vanadium bromoperoxidase mediates the direct transfer of the peroxide oxygen to the sulfide. A schematic model of the vanadium haloperoxidase sulfoxidation mechanism is presented.

Oxidation reactions catalyzed by vanadium chloroperoxidase from Curvularia inaequalis.

Vanadium haloperoxidases have been reported to mediate the oxidation of halides to hypohalous acid and the sulfoxidation of organic sulfides to the corresponding sulfoxides in the presence of hydrogen peroxide. However, traditional heme peroxidase substrates were reported not to be oxidized by vanadium haloperoxidases. Surprisingly, we have now found that the recombinant vanadium chloroperoxidase from the fungus Curvularia inaequalis catalyzes the oxidation of 2,2'-azino-bis(3-ethylbenzthiazoline-6-sulfonic acid) (ABTS), a classical chromogenic heme peroxidase substrate. The enzyme mediates the oxidation of ABTS in the presence of hydrogen peroxide with a turnover frequency of 11 s(-1) at its pH optimum of 4.0. The Km of the recombinant enzyme for ABTS was observed to be approximately 35 microM at this pH value. In addition, the bleaching of an industrial sulfonated azo dye, Chicago Sky Blue 6B, catalyzed by the recombinant vanadium chloroperoxidase in the presence of hydrogen peroxide is reported.

Birefringence of the human crystalline lens in vivo.

We studied aspects of the birefringence in the human crystalline lens. With the use of Mueller-matrix ellipsometry in vivo on both accommodated and unaccommodated eyes, we found no difference between the associated retardations. We calculated form birefringence of the lens by interpreting the membranes of the lens fiber cells as Wiener bodies. The resulting retardation for a light beam that passes the lens as in the experiments exceeds by far the measured total retardation. We conclude that form and intrinsic birefringence of the lens cancel out.

Birefringence of the human foveal area assessed in vivo with Mueller-matrix ellipsometry.

We have assessed retinal birefringence in the foveal and parafoveal regions by applying Mueller matrix ellipsometry on the human eye in vivo. Basically, a light beam passed the ocular media twice and was scattered at the fundus intermediately. Keeping the entry and exit positions on the cornea constant and varying the retinal location along a circle around the foveal center enabled us to separate the corneal and retinal components of the measured retardation. We conclude that the retina within the outer margin of the parafovea behaves as a uniaxial crystal, with its slow axis radially oriented from the fovea and a retardation of about 16 deg (to 70 deg in the corneal center). We believe that Henle's fiber layer causes retardation in this specific configuration of entrance and exit beams. The outer segments of the photoreceptors, although birefringent, have their optic axes aligned with these beams.