title: Glu-370 in the large subunit influences the substrate binding, allosteric, and heat stability properties of potato ADP-glucose pyrophosphorylase.
abstract: ADP-glucose pyrophosphorylase (AGPase) is a key allosteric enzyme in plant starch biosynthesis. Plant AGPase is a heterotetrameric enzyme that consists of large (LS) and small subunits (SS), which are encoded by two different genes. In this study, we showed that the conversion of Glu to Gly at position 370 in the LS of AGPase alters the heterotetrameric stability along with the binding properties of substrate and effectors of the enzyme. Kinetic analyses revealed that the affinity of the LSE370GSSWTAGPase for glucose-1-phosphate is 3-fold less than for wild type (WT) AGPase. Additionally, the LSE370GSSWTAGPase requires 3-fold more 3-phosphogyceric acid to be activated. Finally, the LSE370GSSWTAGPase is less heat stable compared with the WT AGPase. Computational analysis of the mutant Gly-370 in the 3D modeled LS AGPase showed that this residue changes charge distribution of the surface and thus affect stability of the LS AGPase and overall heat stability of the heterotetrameric AGPase. In summary, our results show that LSE370intricately modulate the heat stability and enzymatic activity of potato the AGPase.
authors.affiliation: Koc University, Department of Chemical and Biological Engineering, Rumelifeneri Yolu, Sariyer, Istanbul, Turkey.
authors.forename: Ayse Bengisu
authors.initials: AB
authors.lastname: Seferoglu
authors.affiliation: Koc University, Department of Chemical and Biological Engineering, Rumelifeneri Yolu, Sariyer, Istanbul, Turkey.
authors.forename: Seref
authors.initials: S
authors.lastname: Gul
authors.affiliation: Koc University, Department of Molecular Biology and Genetics, Rumelifeneri Yolu, Sariyer, Istanbul, Turkey.
authors.forename: Ugur Meric
authors.initials: UM
authors.lastname: Dikbas
authors.affiliation: Koc University, Department of Molecular Biology and Genetics, Rumelifeneri Yolu, Sariyer, Istanbul, Turkey.
authors.forename: Ibrahim
authors.initials: I
authors.lastname: Baris
authors.affiliation: Koc University, Department of Chemical and Biological Engineering, Rumelifeneri Yolu, Sariyer, Istanbul, Turkey.
authors.forename: Kaan
authors.initials: K
authors.lastname: Koper
authors.affiliation: Istanbul University, Department of Biology, 34134 Suleymaniye, Istanbul, Turkey.
authors.forename: Mahmut
authors.initials: M
authors.lastname: Caliskan
authors.affiliation: Istanbul University, Department of Biology, 34134 Suleymaniye, Istanbul, Turkey.
authors.forename: Gul
authors.initials: G
authors.lastname: Cevahir
authors.affiliation: Koc University, Department of Chemical and Biological Engineering, Rumelifeneri Yolu, Sariyer, Istanbul, Turkey; Koc University, Department of Molecular Biology and Genetics, Rumelifeneri Yolu, Sariyer, Istanbul, Turkey. Electronic address: hkavakli@ku.edu.tr.
authors.forename: Ibrahim Halil
authors.initials: IH
authors.lastname: Kavakli
mesh_terms: D001665:Binding Sites; D004795:Enzyme Stability; D051599:Glucose-1-Phosphate Adenylyltransferase / Q000737:chemistry / Q000502:physiology*; D006003:Glycogen / Q000096:biosynthesis; D006358:Hot Temperature; D007700:Kinetics; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D010940:Plant Proteins / Q000737:chemistry / Q000502:physiology*; D017434:Protein Structure, Tertiary; D011198:Solanum tuberosum / Q000201:enzymology* / Q000235:genetics; D013213:Starch / Q000096:biosynthesis*; D013379:Substrate Specificity
chemical_list: D010940:Plant Proteins; D013213:Starch; D006003:Glycogen; D051599:Glucose-1-Phosphate Adenylyltransferase
keywords: ADP-glucose pyrophosphorylase; Allosteric regulation; Protein stability; Starch biosynthesis