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Functional analysis of the GTPases EngA and YhbZ encoded by Salmonella typhimurium

Lookup NU author(s): Dr Heather Lamb, Paul Thompson, Dr Nick Watkins, Professor Alastair Hawkins


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The S. typhimurium genome encodes proteins, designated EngA and YhbZ, which have a high sequence identity with the GTPases EngA/Der and ObgE/CgtA E of Escherichia coli. The wild-type activity of the E. coli proteins is essential for normal ribosome maturation and cell viability. In order to characterize the potential involvement of the Salmonella typhimurium EngA and YhbZ proteins in ribosome biology, we used high stringency affinity chromatography experiments to identify strongly binding ribosomal partner proteins. A combination of biochemical and microcalorimetric analysis was then used to characterize these protein:protein interactions and quantify nucleotide binding affinities. These experiments show that YhbZ specifically interacts with the pseudouridine synthase RluD (KD = 2 μM and 1:1 stoichiometry), and we show for the first time that EngA can interact with the ribosomal structural protein S7. Thermodynamic analysis shows both EngA and YhbZ bind GDP with a higher affinity than GTP (20-fold difference for EngA and 3.8-fold for YhbZ), and that the two nucleotide binding sites in EngA show a 5.3-fold difference in affinity for GDP. We report a fluorescence assay for nucleotide binding to EngA and YhbZ, which is suitable for identifying inhibitors specific for this ligand-binding site, which would potentially inhibit their biological functions. The interactions of YhbZ with ribosome structural proteins that we identify may demonstrate a previously unreported additional function for this class of GTPase: that of ensuring delivery of rRNA modifying enzymes to the appropriate region of the ribosome. Published by Cold Spring Harbor Laboratory Press. Copyright © 2007 The Protein Society.

Publication metadata

Author(s): Lamb HK, Thompson P, Elliott C, Charles IG, Richards J, Lockyer M, Watkins N, Nichols C, Stammers DK, Bagshaw CR, Cooper A, Hawkins AR

Publication type: Article

Publication status: Published

Journal: Protein Science

Year: 2007

Volume: 16

Issue: 11

Pages: 2391-2402

ISSN (print): 0961-8368

ISSN (electronic): 1469-896X

Publisher: Wiley-Blackwell Publishing, Inc.


DOI: 10.1110/ps.072900907

PubMed id: 17905831


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Funder referenceFunder name
APG19114Biotechnology and Biological Sciences Research Council