HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

This Article Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Request Permissions Citing Articles Citing Articles via Google Scholar Google Scholar Articles by Casola, L. Articles by Massey, V. Search for Related Content PubMed PubMed Citation Articles by Casola, L. Articles by Massey, V. Social Bookmarking                      What's this?

The Reversible Conversion of Lipoyl Dehydrogenase to an Artifactual Enzyme by Oxidation of Sulfhydryl Groups

From the ¹ From the Department of Biological Chemistry, The University of Michigan, Ann Arbor, Michigan 48104

The effect of cupric ions on the activity, fluorescence, and absorbance, and on the sulfhydryl and flavin adenine dinucleotide contents, of lipoyl dehydrogenase has been examined. The changes take place in two phases during the reaction of the enzyme with copper ions.

In the first phase, the lipoic activity of the enzyme decreases to about 10% of its initial value, while its diaphorase activity increases about 15 times. Diaphorase activity is defined in terms of the enzyme-catalyzed reduction of 2,6-dichloroindophenol at the expense of reduced diphosphopyridine nucleotide. Two sulfhydryl groups are oxidized to a disulfide at this stage, and a slight change in the absorption spectrum occurs, but no change in fluorescence or FAD content takes place. Oxidation of sulfhydryl groups to disulfides continues at a slow rate after the added copper ions have been removed. Changes taking place during this phase of the reaction can be reversed by treatment with cysteine.

Longer exposure of the enzyme to cupric ions leads to a decrease in the diaphorase activity, a decrease in flavin content and fluorescence, and an increase in the fluorescence of aromatic amino acid residues. Further oxidation of sulfhydryl groups occurs which cannot be accounted for entirely by an increase in disulfide content. Changes taking place during this phase of the reaction occur less rapidly at low pH and cannot be reversed by cysteine.

It is concluded that the oxidation of two sulfhydryl groups to a disulfide is responsible for the changes in catalytic properties of the enzyme and that further oxidation results in denaturation, associated with changes in fluorescence and FAD content.

CiteULike    Complore    Connotea    Del.icio.us    Digg    Reddit    Technorati    What's this?