title: Tetramerization of human guanylate-binding protein 1 is mediated by coiled-coil formation of the C-terminal α-helices.
abstract: The human guanylate-binding protein 1 (hGBP1) is a large GTP-binding protein belonging to the dynamin family, a common feature of which is nucleotide-dependent assembly to homotypic oligomers. Assembly leads to stimulation of GTPase activity, which, in the case of dynamin, is responsible for scission of vesicles from membranes. By yeast two-hybrid and biochemical experiments we addressed intermolecular interactions between all subdomains of hGBP1 and identified the C-terminal subdomain, α12/13, as a new interaction site for self-assembly. α12/13 represents a stable subdomain of hGBP1, as shown by CD spectroscopy. In addition to contacts between GTPase domains leading to dimer formation, the interaction between two α12/13 subdomains, in the course of GTP hydrolysis, results in tetramer formation of the protein. With the help of CD spectroscopy we showed coiled-coil formation of two α12/13 subdomains and concentration-dependent measurements allow estimating a value for the dissociation constant of 7.3 μM. We suggest GTP hydrolysis-driven release of the α12/13 subdomain, making it available for coiled-coil formation. Furthermore, we can demonstrate the biological relevance of hGBP1 tetramer formation in living cells by chemical cross-link experiments.
authors.affiliation: Physical Chemistry I, Ruhr-University Bochum, Bochum, Germany.
authors.forename: Adrian
authors.initials: A
authors.lastname: Syguda
authors.forename: Michael
authors.initials: M
authors.lastname: Bauer
authors.forename: Utz
authors.initials: U
authors.lastname: Benscheid
authors.forename: Nicole
authors.initials: N
authors.lastname: Ostler
authors.forename: Elisabeth
authors.initials: E
authors.lastname: Naschberger
authors.forename: Semra
authors.initials: S
authors.lastname: Ince
authors.forename: Michael
authors.initials: M
authors.lastname: Stürzl
authors.forename: Christian
authors.initials: C
authors.lastname: Herrmann
mesh_terms: D001665:Binding Sites / Q000235:genetics; D015153:Blotting, Western; D002478:Cells, Cultured; D002850:Chromatography, Gel; D002942:Circular Dichroism; D019204:GTP-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D006153:Guanosine Diphosphate / Q000737:chemistry / Q000378:metabolism; D006160:Guanosine Triphosphate / Q000737:chemistry / Q000378:metabolism; D061307:Human Umbilical Vein Endothelial Cells / Q000166:cytology / Q000378:metabolism; D006801:Humans; D006868:Hydrolysis; D007700:Kinetics; D008958:Models, Molecular; D011485:Protein Binding; D055503:Protein Multimerization; D020836:Protein Structure, Quaternary; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D020798:Two-Hybrid System Techniques
chemical_list: C490287:GBP1 protein, human; D006153:Guanosine Diphosphate; D006160:Guanosine Triphosphate; D019204:GTP-Binding Proteins