Journal Article

Domain Swapping between a Cyanobacterial and a Plant Subunit ADP-Glucose Pyrophosphorylase

Alberto A. Iglesias, Miguel A. Ballicora, Juliana I. Sesma and Jack Preiss

in Plant and Cell Physiology

Published on behalf of Japanese Society of Plant Physiologists

Volume 47, issue 4, pages 523-530
Published in print April 2006 | ISSN: 0032-0781
Published online April 2006 | e-ISSN: 1471-9053 | DOI: http://dx.doi.org/10.1093/pcp/pcj021
Domain Swapping between a Cyanobacterial and a Plant Subunit ADP-Glucose Pyrophosphorylase

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  • Biochemistry
  • Molecular and Cell Biology
  • Plant Sciences and Forestry

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ADP-glucose pyrophosphorylase (ADP-Glc PPase) catalyzes the regulatory step in the pathway for synthesis of bacterial glycogen and starch in plants. ADP-Glc PPases from cyanobacteria (homotetramer) and from potato (Solanum tuberosum) tuber (heterotetramer) are activated by 3-phosphoglycerate and inhibited by inorganic orthophosphate. To study the function of two putative domains, chimeric enzymes were constructed. PSSANA contained the N-terminus (292 amino acids) of the potato tuber ADP-Glc PPase small subunit (PSS) and the C-terminus (159 residues) of the Anabaena PCC 7120 enzyme. ANAPSS was the inverse chimera. These constructs were expressed separately or together with the large subunit of the potato tuber ADP-Glc PPase (PLS), to obtain homo- and heterotetrameric chimeric proteins. Characterization of these forms showed that the N-terminus determines stability and regulatory redox-dependent properties. The chimeric forms exhibited intermediate 3-phosphoglycerate activation properties with respect to the wild-type homotetrameric enzymes, indicating that the interaction between the putative N- and C-domains determines the affinity for the activator. Characterization of the chimeric heterotetramers showed the functionality of the large subunit, mainly in modulating regulation of the enzyme by the coordinate action of 3-phosphoglycerate and inorganic orthophosphate.

Keywords: ADP-glucose pyrophosphorylase; Cyanobacteria; Glycogen synthesis; Metabolic regulation; Potato tuber; Starch synthesis; ADP-Glc, ADP-glucose; ADP-Glc PPase, ADP-glucose pyrophosphorylase; ANA, C-terminus of Anabaena Pcc 7120 ADP-Glc PPase; BSA, bovine serum albumin; DTT, dithiothreitol; Glc1P, glucose 1-phosphate; 3-PGA, 3-phosphoglycerate; Pi, orthophosphate; PLS, large subunit of the potato tuber ADP-Glc PPase; PPi, pyrophosphate; PSS, small subunit of the potato tuber ADP-Glc PPase

Journal Article.  6831 words.  Illustrated.

Subjects: Biochemistry ; Molecular and Cell Biology ; Plant Sciences and Forestry

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