Journal article
The rice endosperm ADP-glucose pyrophosphorylase large subunit is essential for optimal catalysis and allosteric regulation of the heterotetrameric enzyme
Plant and cell physiology, Vol.55(6), pp.1169-1183
06/2014
Handle:
https://hdl.handle.net/2376/105356
PMID: 24747952
Abstract
Although an alternative pathway has been suggested, the prevailing view is that starch synthesis in cereal endosperm is controlled by the activity of the cytosolic isoform of ADPglucose pyrophosphorylase (AGPase). In rice, the cytosolic AGPase isoform is encoded by the OsAGPS2b and OsAGPL2 genes, which code for the small (S2b) and large (L2) subunits of the heterotetrameric enzyme, respectively. In this study, we isolated several allelic missense and nonsense OsAGPL2 mutants by N-methyl-N-nitrosourea (MNU) treatment of fertilized egg cells and by TILLING (Targeting Induced Local Lesions in Genomes). Interestingly, seeds from three of the missense mutants (two containing T139I and A171V) were severely shriveled and had seed weight and starch content comparable with the shriveled seeds from OsAGPL2 null mutants. Results from kinetic analysis of the purified recombinant enzymes revealed that the catalytic and allosteric regulatory properties of these mutant enzymes were significantly impaired. The missense heterotetramer enzymes and the S2b homotetramer had lower specific (catalytic) activities and affinities for the activator 3-phosphoglycerate (3-PGA). The missense heterotetramer enzymes showed more sensitivity to inhibition by the inhibitor inorganic phosphate (Pi) than the wild-type AGPase, while the S2b homotetramer was profoundly tolerant to Pi inhibition. Thus, our results provide definitive evidence that starch biosynthesis during rice endosperm development is controlled predominantly by the catalytic activity of the cytoplasmic AGPase and its allosteric regulation by the effectors. Moreover, our results show that the L2 subunit is essential for both catalysis and allosteric regulatory properties of the heterotetramer enzyme.
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Details
- Title
- The rice endosperm ADP-glucose pyrophosphorylase large subunit is essential for optimal catalysis and allosteric regulation of the heterotetrameric enzyme
- Creators
- Aytug Tuncel - Institute of Biological Chemistry, Washington State University, Pullman, WA 99164, USAThese authors contributed equally to this workJoe Kawaguchi - Faculty of Agriculture, Kyushu University, Fukuoka, 812-8581 JapanThese authors contributed equally to this workYasuharu Ihara - Faculty of Agriculture, Kyushu University, Fukuoka, 812-8581 JapanHiroaki Matsusaka - Faculty of Agriculture, Kyushu University, Fukuoka, 812-8581 JapanAiko Nishi - Faculty of Agriculture, Kyushu University, Fukuoka, 812-8581 JapanTetsuhiro Nakamura - Faculty of Agriculture, Kyushu University, Fukuoka, 812-8581 JapanSatoru Kuhara - Department of Genetic Resources Technology, Kyushu University, Fukuoka, 812-8581 JapanHideki Hirakawa - Kazusa DNA Research Institute, Department of Plant Genome Research, Kisarazu, JapanYasunori Nakamura - Faculty of Bioresource Sciences, Akita Prefectural University, Akita City, 010-0195 JapanBilal Cakir - Institute of Biological Chemistry, Washington State University, Pullman, WA 99164, USAAi Nagamine - Institute of Biological Chemistry, Washington State University, Pullman, WA 99164, USAFaculty of Agriculture, Kyushu University, Fukuoka, 812-8581 JapanThomas W Okita - Institute of Biological Chemistry, Washington State University, Pullman, WA 99164, USA okita@wsu.eduSeon-Kap Hwang - Institute of Biological Chemistry, Washington State University, Pullman, WA 99164, USAHikaru Satoh - Faculty of Agriculture, Kyushu University, Fukuoka, 812-8581 Japan
- Publication Details
- Plant and cell physiology, Vol.55(6), pp.1169-1183
- Academic Unit
- Biological Chemistry, Institute of
- Publisher
- Japan
- Identifiers
- 99900546862001842
- Language
- English
- Resource Type
- Journal article