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Titolo:
Dimerization of Escherichia coli DNA-gyrase B provides a structural mechanism for activating the ATPase catalytic center
Autore:
Brino, L; Urzhumtsev, A; Mousli, M; Bronner, C; Mitschler, A; Oudet, P; Moras, D;
Indirizzi:
Univ Strasbourg 1, INSERM, CNRS, IGBMC, F-67404 Illkirch, France Univ Strasbourg 1 Illkirch France F-67404 GBMC, F-67404 Illkirch, France Univ Nancy 1, Fac Sci, Lab Cristallog & Modelisat Mat Mineraux & Biol, F-54506 Vandoeuvre Nancy, France Univ Nancy 1 Vandoeuvre Nancy France F-54506 06 Vandoeuvre Nancy, France INSERM, U425, Fac Pharm, F-67401 Illkirch Graffenstaden, France INSERM Illkirch Graffenstaden France F-67401 kirch Graffenstaden, France
Titolo Testata:
JOURNAL OF BIOLOGICAL CHEMISTRY
fascicolo: 13, volume: 275, anno: 2000,
pagine: 9468 - 9475
SICI:
0021-9258(20000331)275:13<9468:DOECDB>2.0.ZU;2-F
Fonte:
ISI
Lingua:
ENG
Soggetto:
ELECTRON-DENSITY MAPS; N-TERMINAL FRAGMENT; TOPOISOMERASE-II; 2-GATE MECHANISM; PROTEIN; MICROSCOPY; NOVOBIOCIN; TRANSPORT; BINDING; MODELS;
Tipo documento:
Article
Natura:
Periodico
Settore Disciplinare:
Life Sciences
Citazioni:
34
Recensione:
Indirizzi per estratti:
Indirizzo: Moras, D Univ Strasbourg 1, INSERM, CNRS, IGBMC, BP 163,1 Rue Laurent Fries, F-67404 Illkirch, France Univ Strasbourg 1 BP 163,1 Rue Laurent Fries Illkirch France F-67404
Citazione:
L. Brino et al., "Dimerization of Escherichia coli DNA-gyrase B provides a structural mechanism for activating the ATPase catalytic center", J BIOL CHEM, 275(13), 2000, pp. 9468-9475

Abstract

DNA-gyrase exhibits an unusual ATP-binding site that is formed as a resultof gyrase B subunit dimerization, a structural transition that is also essential for DNA capture during the topoisomerization cycle, Previous structural studies on Escherichia coli DNA-gyrase B revealed that dimerization is the result of a polypeptidic exchange involving the N-terminal 14 amino acids. To provide experimental data that dimerization is critical for ATPase activity and enzyme turnover, we generated mutants with reduced dimerizationby mutating the two most conserved residues of the GyrB N-terminal arm (Tyr-5 and IIe-10 residues). Our data demonstrate that the hydrophobic IIe-10 residue plays an important role in enzyme dimerization and the nucleotide-protein contact mediated by Tyr-5 side chain residue helps the dimerization process. Analysis of ATPase activities of mutant proteins provides evidencethat dimerization enhances the ATP-hydrolysis turnover. The structure of the Y5S mutant of the N-terminal 43-kDa fragment of E. coli DNA GyrB subunitindicates that Tyr-5 residue provides a scaffold for the ATP-hydrolysis center. We describe a channel formed at the dimer interface that provides a structural mechanism to allow reactive water molecules to access the gamma-phosphate group of the bound ATP molecule. Together, these results demonstrate that dimerization strongly contributes to the folding and stability of the catalytic site for ATP hydrolysis. A role for the essential Mg2+ ion forthe orientation of the phosphate groups of the bound nucleotide inside thereactive pocket was also uncovered by superposition of the 5'-adenylyl beta-gamma-imidodiphosphate (ADPNP) wild-type structure to the salt-free ADPNPstructure.

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Documento generato il 02/10/20 alle ore 02:02:53