Degree Type
Dissertation
Date of Award
1991
Degree Name
Doctor of Philosophy
Department
Biochemistry, Biophysics and Molecular Biology
First Advisor
Carl L. Tipton
Abstract
Two oxysterols, 20(R and S)-20,25-dihydroperoxy-5-cholesten-3[beta]-ol (HPs) isolated from the mixture of cholesterol autoxidation products have been found to reduce effectively cholesteryl ester accumulation in mouse peritoneal macrophages. The effect of HPs was associated with inhibition of cholesterol esterification by suppressing acyl-CoA:cholesterol acyltransferase activity, and by interruption of acetylated low-density-lipoprotein metabolism. The mechanism by which HPs inhibit ACAT activity is investigated in detail in the present study. With reconstituted liposomes, I demonstrated that an increase in cholesterol concentration did not prevent the inhibitory effect of HPs. On the other hand, inactivated ACAT activity was partially restored by treatment with reducing agents, including sodium borohydride, N-methyl-mercaptoacetamide, 2,3-dimercaptopropanol and dithiothreitol. These results suggest that HPs affect the enzyme activity, most likely, through oxidation rather than acting as competitive inhibitors. The reactivities of HPs were studied with model peptides and protein and it was found that the protein sulfhydryl and thioether groups were susceptible to the oxidation by HPs. With specific thiol-modification agent, phenylarsine oxide (PAO), I demonstrated the importance of vicinal sulfhydryl groups for ACAT activity. The facts that phenylarsine oxide-reacted ACAT was insensitive to further inactivation by HP-B and that the same extent of reactivation by 2,3-dimercaptopropanol could be achieved after PAO plus HP-B treatment and after PAO alone suggest that HPs and phenylarsine oxide share common reactive sites. Finally, I proposed a mechanism of two-step modification to explain how HPs inhibit ACAT activity.
DOI
https://doi.org/10.31274/rtd-180813-11677
Publisher
Digital Repository @ Iowa State University, http://lib.dr.iastate.edu/
Copyright Owner
Meiling Shih
Copyright Date
1991
Language
en
Proquest ID
AAI9212188
File Format
application/pdf
File Size
106 pages
Recommended Citation
Shih, Meiling, "Effects of oxysterols on acyl-CoA: cholesterol acyltranferase " (1991). Retrospective Theses and Dissertations. 9685.
https://lib.dr.iastate.edu/rtd/9685