Activity and spectroscopic properties of bovine liver catalase in sodium bis(2-ethylhexyl)sulfosuccinate/isooctane reverse micelles.
Spectroscopic properties such as ultraviolet-visible spectroscopy, circular dichroism, steady-state fluorescence and the catalytic activity of bovine liver catalase (BLC) have been studied in reverse-micelles microemulsions formed by sodium bis(2-ethylhexyl)sulfosuccinate (AOT) in isooctane. Activity measurements were carried out with the highly water soluble hydrogen peroxide as substrate as a function of temperature and pH, at various concentrations of substrate, enzyme, AOT and at different w0 values (w0 = [H2O]/[AOT]). The folding of catalase in reverse micelles is not drastically altered, although there are changes in the local surrounding of the prosthetic group. Kinetic measurements have been carried out under first-order conditions at low substrate concentration. They indicate that the measured substrate decomposition rate is limited by the exchange of substrate molecules between the substrate-filled and the enzyme-containing micelles. The specificity constant, ks = kcat/KM, as related to water pool concentrations, k(s,wp)rm, was, at all w0 values, considerably lower than ks measured in buffer (ksb). In contrast, overall values of ks in reverse micelles, k(s,ov)rm, were always 2-4-times higher than ksb. This apparent superactivity can, however, be ascribed to concentration effects. No 'bell-curve' to describe the w0 dependence of catalase activity in reverse micelles was found, k or ks increasing monotonously up to w0 = 50.
Haber
J
J
Institute of Catalysis and Surface Chemistry, Polish Academy of Sciences.
Maślakiewicz
P
P
Rodakiewicz-Nowak
J
J
Walde
P
P
D000818:Animals; D002374:Catalase / Q000737:chemistry / Q000378:metabolism*; D002384:Catalysis; D002417:Cattle; D002942:Circular Dichroism; D004143:Dioctyl Sulfosuccinic Acid; D006861:Hydrogen Peroxide / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008099:Liver / Q000201:enzymology*; D008823:Micelles; D009795:Octanes; D013050:Spectrometry, Fluorescence; D013056:Spectrophotometry, Ultraviolet; D013696:Temperature
D008823:Micelles; D009795:Octanes; D004143:Dioctyl Sulfosuccinic Acid; D006861:Hydrogen Peroxide; D002374:Catalase; C045798:2,2,4-trimethylpentane