Degree Type


Date of Award


Degree Name

Doctor of Philosophy


Biochemistry, Biophysics and Molecular Biology

First Advisor

James A. Thomas


Mechanisms of S-thiolation and dethiolation of creatine kinase and glycogen phosphorylase b were studied. S-thiolation of the two proteins was initiated by oxyradicals generated by xanthine oxidase in reaction mixtures containing reduced glutathione (GSH). Both proteins were extensively modified under the conditions in which the oxidation of glutathione was inadequate to cause S-thiolation by thiol-disulfide exchange. Creatine kinase was both S-thiolated and non-reducibly oxidized at low GSH concentration, but with adequate GSH, oxidation was prevented while S-thiolation was still active. S-thiolation of glycogen phosphorylase b was not significantly affected by GSH concentration and non-reducible oxidation was not observed. The results suggested that protein sulfhydryls could react more actively than GSH with superoxide anion. Thus, oxyradicals may react with protein sulfhydryls resulting in S-thiolation by a mechanism that involves the reaction of an activated protein thiol with reduced glutathione. Dethiolation of phosphorylase b was actively catalyzed by a cardiac extract by both an NADPH-dependent and a GSH-dependent process. In contrast, creatine kinase was actively dethiolated only by GSH nonenzymatically. Thus, two proteins were dethiolated by different mechanisms. Cells seem to have very high capacity for dethiolation by these three mechanisms. The NADPH-dependent dethiolase had both a high molecular weight component and a low molecular weight component. These two components were sensitive to inhibition by phenylarsine oxide and the inhibition was reversed by addition of a dithiol. Therefore, the properties of these components were similar to thioredoxin and thioredoxin reductase. Dethiolation of phosphorylase b was also demonstrated by Escherichia coli thioredoxin with either dithiothreitol or cardiac extract and NADPH as a reducing power, while creatine kinase was not dethiolated by this process. In contrast, GSH-dependent dethiolation required a single component of low molecular weight, and it was less sensitive to phenylarsine oxide inhibition;Therefore, protein S-thiolation and dethiolation play an important role in preventing oxidative damage to cellular system during oxidative stress.



Digital Repository @ Iowa State University,

Copyright Owner

Eun-Mi Choi Park



Proquest ID


File Format


File Size

145 pages

Included in

Biochemistry Commons