Mechanism of reductive activation of potato tuber ADP-glucose pyrophosphorylase

Yingbin Fu, Miguel A. Ballicora, Joseph F. Leykam, Jack Preiss

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117 Scopus citations


The potato tuber (Solanum tuberosum L.) ADP-glucose pyrophosphorylase activity is activated by a incubation with ADP-glucose and dithiothreitol or by ATP, glucose- 1-phosphate, Ca2+, and dithiothreitol. The activation was accompanied by the appearance of new sulfhydryl groups as determined with 5,5'-dithiobis(2-nitrobenzoic acid). By analyzing the activated and nonactivated enzymes on SDS-polyacrylamide gel electrophoresis under nonreducing conditions, it was found that an intermolecular disulfide bridge between the small subunits of the potato tuber enzyme was reduced during the activation. Further experiments showed that the activation was mediated via a slow reduction and subsequent rapid conformational change induced by ADP- glucose. The activation process could be reversed by oxidation with 5,5'- dithiobis(2-nitrobenzoic acid). Incubation with ADP-glucose and dithiothreitol could reactivate the oxidized enzyme. Chemical modification experiments with [14C]iodoacetic acid and 4-vinylpyridine determined that the intermolecular disulfide bridge was located between Cys12 of the small subunits of the potato tuber enzyme. Mutation of Cys12 in the small subunit into either Ala or Ser eliminated the requirement of DTT on the activation and prevented the formation of the intermolecular disulfide of the potato tuber enzyme. The mutants had instantaneous activation rates as the wild- type in the reduced state. A two-step activation model is proposed.

Original languageEnglish (US)
Pages (from-to)25045-25052
Number of pages8
JournalJournal of Biological Chemistry
Issue number39
StatePublished - Sep 25 1998
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry


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