title: Molecular landscape of the interaction between the urease accessory proteins UreE and UreG.
abstract: Urease, the most efficient enzyme so far discovered, depends on the presence of nickel ions in the catalytic site for its activity. The transformation of inactive apo-urease into active holo-urease requires the insertion of two Ni(II) ions in the substrate binding site, a process that involves the interaction of four accessory proteins named UreD, UreF, UreG and UreE. This study, carried out using calorimetric and NMR-based structural analysis, is focused on the interaction between UreE and UreG from Sporosarcina pasteurii, a highly ureolytic bacterium. Isothermal calorimetric protein-protein titrations revealed the occurrence of a binding event between SpUreE and SpUreG, entailing two independent steps with positive cooperativity (Kd1=42±9μM; Kd2=1.7±0.3μM). This was interpreted as indicating the formation of the (UreE)2(UreG)2 hetero-oligomer upon binding of two UreG monomers onto the pre-formed UreE dimer. The molecular details of this interaction were elucidated using high-resolution NMR spectroscopy. The occurrence of SpUreE chemical shift perturbations upon addition of SpUreG was investigated and analyzed to establish the protein-protein interaction site. The latter appears to involve the Ni(II) binding site as well as mobile portions on the C-terminal and the N-terminal domains. Docking calculations based on the information obtained from NMR provided a structural basis for the protein-protein contact site. The high sequence and structural similarity within these protein classes suggests a generality of the interaction mode among homologous proteins. The implications of these results on the molecular details of the urease activation process are considered and analyzed.
authors: Laboratory of Bioinorganic Chemistry, Department of Pharmacy and Biotechnology, University of Bologna, Italy.
authors: Anna
authors: A
authors: Merloni
authors: Laboratory of Bioinorganic Chemistry, Department of Pharmacy and Biotechnology, University of Bologna, Italy.
authors: Olena
authors: O
authors: Dobrovolska
authors: Laboratory of Bioinorganic Chemistry, Department of Pharmacy and Biotechnology, University of Bologna, Italy.
authors: Barbara
authors: B
authors: Zambelli
authors: Laboratory of Bioinorganic Chemistry, Department of Pharmacy and Biotechnology, University of Bologna, Italy.
authors: Federico
authors: F
authors: Agostini
authors: Laboratory of Bioinorganic Chemistry, Department of Pharmacy and Biotechnology, University of Bologna, Italy.
authors: Micaela
authors: M
authors: Bazzani
authors: Laboratory of Bioinorganic Chemistry, Department of Pharmacy and Biotechnology, University of Bologna, Italy.
authors: Francesco
authors: F
authors: Musiani
authors: Laboratory of Bioinorganic Chemistry, Department of Pharmacy and Biotechnology, University of Bologna, Italy. Electronic address: stefano.ciurli@unibo.it.
authors: Stefano
authors: S
authors: Ciurli
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D002151:Calorimetry; D002352:Carrier Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D002413:Cations, Divalent; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D062105:Molecular Docking Simulation; D009532:Nickel / Q000737:chemistry* / Q000378:metabolism; D028044:Phosphate-Binding Proteins; D011485:Protein Binding; D054730:Protein Interaction Domains and Motifs; D055503:Protein Multimerization; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D058644:Sporosarcina / Q000737:chemistry* / Q000201:enzymology; D013816:Thermodynamics; D014510:Urease / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D002352:Carrier Proteins; D002413:Cations, Divalent; D028044:Phosphate-Binding Proteins; D011994:Recombinant Proteins; C090404:UreE protein, Bacteria; C082270:ureG protein, Bacteria; D009532:Nickel; D014510:Urease
keywords: Calorimetry; Nuclear magnetic resonance; Protein–protein docking; UreE; UreG; Urease