Modulation of Allosteric Regulation by E38K and G101N Mutations in the Potato Tuber ADP-glucose Pyrophosphorylase

Wakuta, Shinji; Shibata, Y.; Yoshizaki, Yumiko; Saburi, Wataru; Hamada, Shigeki; Ito, Hiroyuki; Hwang, Seon‐Kap; Okita, Thomas W.; Matsui, Hirokazu

doi:10.1271/bbb.130276

Cited by 3 publications

(2 citation statements)

References 36 publications

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“…Fructose 2,6-bisphosphate (Fru2,6BP) and PEP are only partially effective (<30 % of levels obtained with 3-PGA) in activating the enzyme, while Fru6P and Glc6P are very weak activators (Iglesias et al 1993;Wakuta et al 2013). The potato AGPase is activated more than 30-fold by 3-PGA and inhibited by Pi.…”

Section: Sink Enzymesmentioning

confidence: 99%

Starch

2015

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Section: Sink Enzymesmentioning

confidence: 99%

Starch

2015

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“…Therefore, it is very important for the regulation of activity of AGPase. To date, there are three main ways reported to regulate the activity of AGPase: small effector molecules for allosteric regulation, thermal regulation and redox regulation [9,10,11,12,38]. In this study, we used phos-tag TM technology, Pro-Q diamond staining and iTRAQ TM to investigate phosphorylation of AGPase.…”

Section: Discussionmentioning

confidence: 99%

The Proteomic Analysis of Maize Endosperm Protein Enriched by Phos-tagtm Reveals the Phosphorylation of Brittle-2 Subunit of ADP-Glc Pyrophosphorylase in Starch Biosynthesis Process

Shen

et al. 2019

IJMS

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AGPase catalyzes a key rate-limiting step that converts ATP and Glc-1-p into ADP-glucose and diphosphate in maize starch biosynthesis. Previous studies suggest that AGPase is modulated by redox, thermal and allosteric regulation. However, the phosphorylation of AGPase is unclear in the kernel starch biosynthesis process. Phos-tagTM technology is a novel method using phos-tagTM agarose beads for separation, purification, and detection of phosphorylated proteins. Here we identified phos-tagTM agarose binding proteins from maize endosperm. Results showed a total of 1733 proteins identified from 10,678 distinct peptides. Interestingly, a total of 21 unique peptides for AGPase sub-unit Brittle-2 (Bt2) were identified. Bt2 was demonstrated by immunoblot when enriched maize endosperm protein with phos-tagTM agarose was in different pollination stages. In contrast, Bt2 would lose binding to phos-tagTM when samples were treated with alkaline phosphatase (ALP). Furthermore, Bt2 could be detected by Pro-Q diamond staining specifically for phosphorylated protein. We further identified the phosphorylation sites of Bt2 at Ser10, Thr451, and Thr462 by iTRAQ. In addition, dephosphorylation of Bt2 decreased the activity of AGPase in the native gel assay through ALP treatment. Taking together, these results strongly suggest that the phosphorylation of AGPase may be a new model to regulate AGPase activity in the starch biosynthesis process.

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