PMID:9677420
| Citation |
Wower, IK, Wower, J and Zimmermann, RA (1998) Ribosomal protein L27 participates in both 50 S subunit assembly and the peptidyl transferase reaction. J. Biol. Chem. 273:19847-52 |
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| Abstract |
Protein L27 has been implicated as a constituent of the peptidyl transferase center of the Escherichia coli 50 S ribosomal subunit by a variety of experimental observations. To define better the functional role of this protein, we constructed a strain in which the rpmA gene, which encodes L27, was replaced by a kanamycin resistance marker. The deletion mutant grows five to six times slower than the wild-type parent and is both cold- and temperature-sensitive. This phenotype is reversed when L27 is expressed from a plasmid-borne copy of the rpmA gene. Analysis of ribosomes from the L27-lacking strain revealed deficiencies in both the assembly and activity of the 50 S ribosomal subunits. Although functional 50 S subunits are formed in the mutant, an assembly "bottleneck" was evidenced by the accumulation of a prominent 40 S precursor to the 50 S subunit which was deficient in proteins L16, L20, and L21, as well as L27. In addition, the peptidyl transferase activity of 70 S ribosomes containing mutant 50 S subunits was determined to be three to four times lower than for wild-type ribosomes. Ribosomes lacking L27 were found to be impaired in the enzymatic binding of Phe-tRNAPhe to the A site, although the interaction of N-acetyl-Phe-tRNAPhe with the P site was largely unperturbed. We therefore infer that L27 contributes to peptide bond formation by facilitating the proper placement of the acceptor end of the A-site tRNA at the peptidyl transferase center. |
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| Keywords |
Binding Sites/genetics; Cross-Linking Reagents/metabolism; Electrophoresis, Gel, Two-Dimensional; Escherichia coli/metabolism; Genetic Markers; Kanamycin/pharmacology; Kinetics; Mutation/genetics; Peptide Elongation Factor Tu/metabolism; Peptidyl Transferases/metabolism; Phenotype; RNA, Transfer, Phe/metabolism; Ribosomal Proteins/physiology; Ribosomes/chemistry |
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