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          Institute: MPI für molekulare Genetik     Collection: Ribosome Research Groups     Display Documents



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ID: 305598.0, MPI für molekulare Genetik / Ribosome Research Groups
EF-G-dependent GTPase on the ribosome: Conformational change and fusidic acid inhibition
Authors:Seo, Hyuk-Soo; Abedin, Sameem; Kamp, Detlev; Wilson, Daniel N.; Nierhaus, Knud; Cooperman, Barry S.
Language:English
Date of Publication (YYYY-MM-DD):2006-02-01
Title of Journal:Biochemistry
Volume:45
Issue / Number:8
Start Page:2504
End Page:2514
Copyright:© 2006 American Chemical Society
Review Status:not specified
Audience:Experts Only
Abstract / Description:Protein synthesis studies increasingly focus on delineating the nature of conformational changes occurring as the ribosome exerts its catalytic functions. Here, we use FRET to examine such changes during single-turnover EF-G-dependent GTPase on vacant ribosomes and to elucidate the mechanism by which fusidic acid (FA) inhibits multiple-turnover EF-G·GTPase. Our measurements focus on the distance between the G' region of EF-G and the N-terminal region of L11 (L11-NTD), located within the GTPase activation center of the ribosome. We demonstrate that single-turnover ribosome-dependent EF-G GTPase proceeds according to a kinetic scheme in which rapid G' to L11-NTD movement requires prior GTP hydrolysis and, via branching pathways, either precedes Pi release (major pathway) or occurs simultaneously with it (minor pathway). Such movement retards Pi release, with the result that Pi release is essentially rate-determining in single-turnover GTPase. This is the most significant difference between the EF-G·GTPase activities of vacant and translocating ribosomes [Savelsbergh, A., Katunin, V. I., Mohr, D., Peske, F., Rodnina, M. V., and Wintermeyer, W. (2003) Mol. Cell 11, 1517-1523], which are otherwise quite similar. Both the G' to L11-NTD movement and Pi release are strongly inhibited by thiostrepton but not by FA. Contrary to the standard view that FA permits only a single round of GTP hydrolysis [Bodley, J. W., Zieve, F. J., and Lin, L. (1970) J. Biol. Chem. 245, 5662-5667], we find that FA functions rather as a slow inhibitor of EF-G·GTPase, permitting a number of GTPase turnovers prior to complete inhibition while inducing a closer approach of EF-G to the GAC than is seen during normal turnover.
External Publication Status:published
Document Type:Article
Communicated by:Ribosome Group
Affiliations:MPI für molekulare Genetik
External Affiliations:Department of Chemistry, University of Pennsylvania, Philadelphia, Pennsylvania 19104-6323, USA.
Identifiers:ISSN:0006-2960
DOI:10.1021/bi0516677
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