Mutagenesis of the glucose-1-phosphate-binding site of potato tuber ADP-glucose pyrophosphorylase

Yingbin Fu, Miguel A. Ballicora, Jack Preiss

Research output: Contribution to journalArticle

Abstract

Lysine (Lys)-195 in the homotetrameric ADP-glucose pyrophosphorylase (ADPGIc PPase) from Escherichia coli was shown previously to be involved in the binding of the substrate glucose-1-phosphate (Glc-1-P). This residue is highly conserved in the ADPGIc PPase family. Site-directed mutagenesis was used to investigate the function of this conserved Lys residue in the large and small subunits of the heterotetrameric potato (Solanum tuberosum) tuber enzyme. The apparent affinity for Glc-1-P of the wild-type enzyme decreased 135- to 550-fold by changing Lys-198 of the small subunit to arginine, alanine, or glutamic acid, suggesting that both the charge and the size of this residue influence Glc-1-P binding. These mutations had little effect on the kinetic constants for the other substrates (ATP and Mg2+ or ADP-GIc and inorganic phosphate), activator (3-phosphoglycerate), inhibitor (inorganic phosphate), or on the thermal stability. Mutagenesis of the corresponding Lys (Lys-213) in the large subunit had no effect on the apparent affinity for Glc-1-P by substitution with arginine, alanine, or glutamic acid. A double mutant, SK198RLK213R, was also obtained that had a 100 fold reduction of the apparent affinity for Glc-1 -P. The data indicate that Lys-198 in the small subunit is directly involved in the binding of Glc-1-P, whereas they appear to exclude a direct role of Lys-213 in the large subunit in the interaction with this substrate.

Original languageEnglish (US)
Pages (from-to)989-996
Number of pages8
JournalPlant Physiology
Volume117
Issue number3
StatePublished - 1998
Externally publishedYes

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Glucose-1-Phosphate Adenylyltransferase
glucose 1-phosphate
Solanum tuberosum
mutagenesis
Mutagenesis
Lysine
binding sites
lysine
tubers
Binding Sites
potatoes
glutamic acid
Alanine
alanine
arginine
Arginine
Glutamic Acid
Phosphates
phosphates
phosphoglyceric acids

ASJC Scopus subject areas

  • Plant Science

Cite this

Mutagenesis of the glucose-1-phosphate-binding site of potato tuber ADP-glucose pyrophosphorylase. / Fu, Yingbin; Ballicora, Miguel A.; Preiss, Jack.

In: Plant Physiology, Vol. 117, No. 3, 1998, p. 989-996.

Research output: Contribution to journalArticle

Fu, Yingbin ; Ballicora, Miguel A. ; Preiss, Jack. / Mutagenesis of the glucose-1-phosphate-binding site of potato tuber ADP-glucose pyrophosphorylase. In: Plant Physiology. 1998 ; Vol. 117, No. 3. pp. 989-996.
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