The Escherichia coli GTPase CgtAE is involved in late steps of large ribosome assembly.
The bacterial ribosome is an extremely complicated macromolecular complex the in vivo biogenesis of which is poorly understood. Although several bona fide assembly factors have been identified, their precise functions and temporal relationships are not clearly defined. Here we describe the involvement of an Escherichia coli GTPase, CgtA(E), in late steps of large ribosomal subunit biogenesis. CgtA(E) belongs to the Obg/CgtA GTPase subfamily, whose highly conserved members are predominantly involved in ribosome function. Mutations in CgtA(E) cause both polysome and rRNA processing defects; small- and large-subunit precursor rRNAs accumulate in a cgtA(E) mutant. In this study we apply a new semiquantitative proteomic approach to show that CgtA(E) is required for optimal incorporation of certain late-assembly ribosomal proteins into the large ribosomal subunit. Moreover, we demonstrate the interaction with the 50S ribosomal subunits of specific nonribosomal proteins (including heretofore uncharacterized proteins) and define possible temporal relationships between these proteins and CgtA(E). We also show that purified CgtA(E) associates with purified ribosomal particles in the GTP-bound form. Finally, CgtA(E) cofractionates with the mature 50S but not with intermediate particles accumulated in other large ribosome assembly mutants.
Jiang
Mengxi
M
Department of Molecular, Cellular and Developmental Biology, University of Michigan, 830 North University, Ann Arbor, MI 48109-1048, USA.
Datta
Kaustuv
K
Walker
Angela
A
Strahler
John
J
Bagamasbad
Pia
P
Andrews
Philip C
PC
Maddock
Janine R
JR
D019943:Amino Acid Substitution / Q000235:genetics; D002458:Cell Fractionation; D053487:DEAD-box RNA Helicases; D004926:Escherichia coli / Q000235:genetics / Q000502:physiology*; D029968:Escherichia coli Proteins / Q000235:genetics / Q000502:physiology*; D020558:GTP Phosphohydrolases / Q000032:analysis / Q000235:genetics / Q000502:physiology*; D017353:Gene Deletion; D020559:Monomeric GTP-Binding Proteins / Q000235:genetics / Q000502:physiology*; D020125:Mutation, Missense; D011485:Protein Binding; D020365:RNA Helicases / Q000235:genetics; D012269:Ribosomal Proteins / Q000378:metabolism*; D012270:Ribosomes / Q000378:metabolism*
D029968:Escherichia coli Proteins; D012269:Ribosomal Proteins; D020558:GTP Phosphohydrolases; C491572:ObgE protein, E coli; C088840:deaD protein, E coli; D053487:DEAD-box RNA Helicases; D020365:RNA Helicases; D020559:Monomeric GTP-Binding Proteins