Analysis of the inactivation of liver alcohol dehydrogenase during storage in Aerosol-OT/isooctane microemulsions.
Changes in the enzymatic properties of horse liver alcohol dehydrogenase (HLADH; EC 1.1.1.1) were studied as a function of incubation time in Aerosol-OT/isooctane microemulsions. The enzyme was characterized by fluorimetric binding studies of the inhibitor isobutyramide to the binary complex, HLADH-NADH and by determination of Km,app and Vmax,app values for cyclohexanone. The Km,app values for cyclohexanone and the Kd,app for isobutyramide stay constant throughout a 48-h incubation, whereas the Vmax,app and the total number of inhibitor binding sites decrease. Thus the inactivation process previously described corresponds to progressive loss of functional sites, while the properties of the remaining functional sites are unchanged. If no co-enzyme is added to the system, the enzyme loses catalytic activity within less than an hour, but if co-enzyme is added, a fraction of the HLADH enzyme population retains enzyme activity over a long period of time. Hence the presence of bound co-enzyme significantly inhibits the process(es) leading to inactivation of the enzyme in the microemulsions.
Larsson
K M
KM
Department of Biotechnology, Chemical Center, University of Lund, Sweden.
Oldfield
C
C
Freedman
R B
RB
D000336:Aerosols; D000426:Alcohol Dehydrogenase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D000577:Amides / Q000378:metabolism; D003512:Cyclohexanones / Q000378:metabolism; D004143:Dioctyl Sulfosuccinic Acid; D004355:Drug Stability; D004655:Emulsions; D007700:Kinetics; D008099:Liver / Q000201:enzymology*; D009243:NAD / Q000378:metabolism; D009795:Octanes; D013050:Spectrometry, Fluorescence; D013501:Surface-Active Agents
D000336:Aerosols; D000577:Amides; D003512:Cyclohexanones; D004655:Emulsions; D009795:Octanes; D013501:Surface-Active Agents; D009243:NAD; D004143:Dioctyl Sulfosuccinic Acid; C036468:cyclohexanone; C030197:isobutyramide; D000426:Alcohol Dehydrogenase; C045798:2,2,4-trimethylpentane