Activation of heterotrimeric G proteins by a high energy phosphate transfer via nucleoside diphosphate kinase (NDPK) B and Gβ subunits

• Verfasst von: Cuello, Friederike [VerfasserIn]
• Verfasst von: Schulze, Rüdiger A. [VerfasserIn]
• Verfasst von: Heemeyer, Frank [VerfasserIn]
• Verfasst von: Meyer, Helmut E. [VerfasserIn]
• Verfasst von: Lutz, Susanne [VerfasserIn]
• Verfasst von: Jakobs, Karl-Heinz [VerfasserIn]
• Verfasst von: Niroomand, Feraydoon [VerfasserIn]
• Verfasst von: Wieland, Thomas [VerfasserIn]
• Titel: Activation of heterotrimeric G proteins by a high energy phosphate transfer via nucleoside diphosphate kinase (NDPK) B and Gβ subunits
• Titelzusatz: complex formation of NDPK B with Gβγ dimers and phosphorylation of His-266 IN Gβ
• Verf.angabe: Friederike Cuello, Rüdiger A. Schulze, Frank Heemeyer, Helmut E. Meyer, Susanne Lutz, Karl H. Jakobs, Feraydoon Niroomand, and Thomas Wieland
• Jahr: 2003
• Umfang: 7 S.
• Fussnoten: Published in Press, December 16, 2002 ; Gesehen am 07.04.2022
• Titel Quelle: Enthalten in: The journal of biological chemistry
• Ort Quelle: Bethesda, Md. : ASBMB Publications, 1905
• Jahr Quelle: 2003
• Band/Heft Quelle: 278(2003), 9, Seite 7220-7226
• ISSN Quelle: 1083-351X
• DOI: doi:10.1074/jbc.M210304200
• Datenträger: Online-Ressource
• Sprache: eng
• Sach-SW: Animals
• Sach-SW: Blotting, Western
• Sach-SW: Brain
• Sach-SW: Cattle
• Sach-SW: Cell Membrane
• Sach-SW: Diethyl Pyrocarbonate
• Sach-SW: Dimerization
• Sach-SW: Dose-Response Relationship, Drug
• Sach-SW: Durapatite
• Sach-SW: GTP-Binding Proteins
• Sach-SW: Humans
• Sach-SW: Immunoblotting
• Sach-SW: Models, Molecular
• Sach-SW: Nucleoside-Diphosphate Kinase
• Sach-SW: Peptides
• Sach-SW: Phosphates
• Sach-SW: Phosphorylation
• Sach-SW: Precipitin Tests
• Sach-SW: Protein Binding
• Sach-SW: Protein Conformation
• Sach-SW: Protein Isoforms
• Sach-SW: Protein Structure, Tertiary
• Sach-SW: Recombinant Proteins
• Sach-SW: Retina
• Sach-SW: Rod Cell Outer Segment
• Sach-SW: Serine Endopeptidases
• Sach-SW: Trypsin
• K10plus-PPN: 1798102625

Abstract

G protein betagamma dimers can be phosphorylated in membranes from various tissues by GTP at a histidine residue in the beta subunit. The phosphate is high energetic and can be transferred onto GDP leading to formation of GTP. Purified Gbetagamma dimers do not display autophosphorylation, indicating the involvement of a separate protein kinase. We therefore enriched the Gbeta-phosphorylating activity present in preparations of the retinal G protein transducin and in partially purified G(i/o) proteins from bovine brain. Immunoblots, autophosphorylation, and enzymatic activity measurements demonstrated enriched nucleoside diphosphate kinase (NDPK) B in both preparations, together with residual Gbetagamma dimers. In the retinal NDPK B-enriched fractions, a Gbeta-specific antiserum co-precipitated phosphorylated NDPK B, and an antiserum against the human NDPK co-precipitated phosphorylated Gbetagamma. In addition, the NDPK-containing fractions from bovine brain reconstituted the phosphorylation of purified Gbetagamma. For identification of the phosphorylated histidine residue, bovine brain Gbetagamma and G(t)betagamma were thiophosphorylated with guanosine 5'-O-(3-[(35)S]thio)triphosphate, followed by digestion with endoproteinase Glu-C and trypsin, separation of the resulting peptides by gel electrophoresis and high pressure liquid chromatography, respectively, and sequencing of the radioactive peptides. The sequence information produced by both methods identified specific labeled fragments of bovine Gbeta(1) that overlapped in the heptapeptide, Leu-Met-Thr-Tyr-Ser-His-Asp (amino acids 261-267). We conclude that NDPK B forms complexes with Gbetagamma dimers and contributes to G protein activation by increasing the high energetic phosphate transfer onto GDP via intermediately phosphorylated His-266 in Gbeta(1) subunits.