title: Ribosome engineering to promote new crystal forms.
abstract: Crystallographic studies of the ribosome have provided molecular details of protein synthesis. However, the crystallization of functional complexes of ribosomes with GTPase translation factors proved to be elusive for a decade after the first ribosome structures were determined. Analysis of the packing in different 70S ribosome crystal forms revealed that regardless of the species or space group, a contact between ribosomal protein L9 from the large subunit and 16S rRNA in the shoulder of a neighbouring small subunit in the crystal lattice competes with the binding of GTPase elongation factors to this region of 16S rRNA. To prevent the formation of this preferred crystal contact, a mutant strain of Thermus thermophilus, HB8-MRCMSAW1, in which the ribosomal protein L9 gene has been truncated was constructed by homologous recombination. Mutant 70S ribosomes were used to crystallize and solve the structure of the ribosome with EF-G, GDP and fusidic acid in a previously unobserved crystal form. Subsequent work has shown the usefulness of this strain for crystallization of the ribosome with other GTPase factors.
authors.affiliation: Department of Cell and Molecular Biology, Uppsala University, Box 596, SE-751 24 Uppsala, Sweden. maria.selmer@icm.uu.se
authors.forename: Maria
authors.initials: M
authors.lastname: Selmer
authors.forename: Yong-Gui
authors.initials: YG
authors.lastname: Gao
authors.forename: Albert
authors.initials: A
authors.lastname: Weixlbaumer
authors.forename: V
authors.initials: V
authors.lastname: Ramakrishnan
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D018360:Crystallography, X-Ray; D005672:Fusidic Acid / Q000737:chemistry / Q000378:metabolism; D006153:Guanosine Diphosphate / Q000737:chemistry / Q000378:metabolism; D059765:Homologous Recombination; D008958:Models, Molecular; D009154:Mutation; D020653:Peptide Elongation Factor G / Q000737:chemistry / Q000378:metabolism; D012269:Ribosomal Proteins / Q000235:genetics; D054681:Ribosome Subunits, Large, Bacterial / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D016963:Thermus thermophilus / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D020653:Peptide Elongation Factor G; D012269:Ribosomal Proteins; C049650:ribosomal protein L9; D006153:Guanosine Diphosphate; D005672:Fusidic Acid