Skip to main content
Article thumbnail
Location of Repository

Distinct GDP/GTP bound states of the tandem G-domains of EngA regulate ribosome binding

By Sushil Kumar Tomar, Neha Dhimole, Moon Chatterjee and Balaji Prakash

Abstract

EngA, a unique GTPase containing a KH-domain preceded by two consecutive G-domains, displays distinct nucleotide binding and hydrolysis activities. So far, Escherichia coli EngA is reported to bind the 50S ribosomal subunit in the guanosine-5′-trihosphate (GTP) bound state. Here, for the first time, using mutations that allow isolating the activities of the two G-domains, GD1 and GD2, we show that apart from 50S, EngA also binds the 30S and 70S subunits. We identify that the key requirement for any EngA–ribosome association is GTP binding to GD2. In this state, EngA displays a weak 50S association, which is further stabilized when GD1 too binds GTP. Exchanging bound GTP with guanosine-5′-diphosphate (GDP), at GD1, results in interactions with 50S, 30S and 70S. Therefore, it appears that GD1 employs GTP hydrolysis as a means to regulate the differential specificity of EngA to either 50S alone or to 50S, 30S and 70S subunits. Furthermore, using constructs lacking either GD1 or both GD1 and GD2, we infer that GD1, when bound to GTP and GDP, adopts distinct conformations to mask or unmask the 30S binding site on EngA. Our results suggest a model where distinct nucleotide-bound states of the two G-domains regulate formation of specific EngA–ribosome complexes

Topics: Molecular Biology
Publisher: Oxford University Press
OAI identifier: oai:pubmedcentral.nih.gov:2673443
Provided by: PubMed Central
Download PDF:
Sorry, we are unable to provide the full text but you may find it at the following location(s):
  • http://www.pubmedcentral.nih.g... (external link)
  • Suggested articles

    Citations

    1. (1988). A malachite green procedure for orthophosphate determination and its use in alkaline phosphatase-based enzyme immunoassay.
    2. (2001). An essential GTPase, Der, containing double GTP-binding domains from Escherichia coli and Thermotoga maritime.
    3. (2001). Analysis of the interaction of 16S rRNA and cytoplasmic membrane with the C-terminal part of the Streptococcus pneumoniae Era GTPase.
    4. (2005). Characterization of the Bacillus subtilis GTPase YloQ and its role in ribosome function.
    5. (2004). Coot: model-building tools for molecular graphics.
    6. (2002). Domain arrangement of Der, a switch protein containing two GTPase domains.
    7. (2008). E. coli HflX interacts with 50S ribosomal subunits in presence of nucleotides.
    8. (1999). Era, an essential Escherichia coli small G-protein, binds to the 30S ribosomal subunit.
    9. (1996). Escherichia coli trigger factor is a prolyl isomerase that associates with nascent polypeptide chains.
    10. (1994). Fluorescent guanine nucleotide analogs and G protein activation.
    11. (2003). Function of the universally conserved bacterial GTPases.
    12. (1997). G protein mechanisms: insights from structural analysis.
    13. (2002). GTPase activation of elongation factors
    14. (2003). Induced nucleotide specificity in a GTPase.
    15. (2006). Multiple GTPases participate in the assembly of the large ribosomal subunit in Bacillus subtilis.
    16. (1993). Mutations in human dynamin block an intermediate stage in coated vesicle formation.
    17. (2002). Overexpression of two different GTPases rescues a null mutation in a heat-induced rRNA methyltransferase.
    18. (2008). Salmonella enterica serovar Typhimurium BipA exhibits two distinct ribosome binding modes.
    19. (2006). Structural stabilization of GTP-binding domains in circularly permuted GTPases: implications for RNA binding.
    20. (2004). Studies of the interaction of Escherichia coli YjeQ with the ribosome in vitro.
    21. (2006). The essential GTPase RbgA (YlqF) is required for 50S ribosome assembly in Bacillus subtilis.
    22. (2006). The essential GTPase YphC displays a major domain rearrangement associated with nucleotide binding.
    23. (2007). The essential GTPase YqeH is required for proper ribosome assembly in Bacillus subtilis.
    24. (2005). The GTP binding protein Obg homolog ObgE is involved in ribosome maturation.
    25. (1990). The GTPase superfamily: a conserved switch for diverse cell functions.
    26. (1991). The GTPase superfamily: conserved structure and molecular mechanism.
    27. (2006). The tandem GTPase, Der, is essential for the biogenesis of 50S ribosomal subunits in Escherichia coli.
    28. (2004). UCSF Chimera–a visualization system for exploratory research and analysis.

    To submit an update or takedown request for this paper, please submit an Update/Correction/Removal Request.