Identification of a region of Escherichia coli ribosomal protein L2 required for the assembly of L16 into the 50 S ribosomal subunit.
In vitro mutagenesis of rplB was used to generate changes in a conserved region of Escherichia coli ribosomal protein L2 between Gly221 and His231. Mutants were selected by temperature sensitivity using an inducible expression system. A mutant L2 protein with the deletion of Thr222 to Asp228 was readily distinguishable from wild-type L2 by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and ribosomes from the strain overexpressing this mutant protein were characterized by sucrose density gradient centrifugation and protein composition. In addition to 30 S and 50 S ribosomal subunits, cell lysates contained a new component that sedimented at 40 S in 1 mM Mg2+ and at 48 S in 10 mM Mg2+. These particles contained mutant L2 protein exclusively, completely lacked L16, and had reduced amounts of L28, L33, and L34. They did not reassociate with 30 S ribosomal subunits and were inactive in polyphenylalanine synthesis. Other mutants in the same conserved region, including the substitution of His229 by Gln229, produced similar aberrant 50 S particles that sedimented at 40 S and failed to associate with 30 S subunits.
Department of Biological Chemistry, School of Medicine, University of California, Davis 95521.
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Romero
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Arredondo
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Traut
D000595:Amino Acid Sequence; D002499:Centrifugation, Density Gradient; D003001:Cloning, Molecular; D004252:DNA Mutational Analysis; D015180:Electrophoresis, Gel, Two-Dimensional; D004926:Escherichia coli; D008969:Molecular Sequence Data; D009024:Morphogenesis; D014176:Protein Biosynthesis; D012269:Ribosomal Proteins / Q000235:genetics / Q000378:metabolism*; D012270:Ribosomes / Q000502:physiology / Q000648:ultrastructure*
D012269:Ribosomal Proteins; C027937:ribosomal protein L16; C029466:ribosomal protein L2