The properties of the complex between ribosomal protein L2 and tRNA

Rèmme, Jaanus; Metspalu, Ene; Maimets, Toivo; Villems, Richard

doi:10.1016/0014-5793(85)81299-0

Cited by 8 publications

(2 citation statements)

References 17 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…More specifically, L16 may be instrumental in catalysis and the binding of aminoacyl-tRNA to the A-site of the ribosome. Similarly ribosomal protein L2, another essential component of peptidyltransferase which has also been shown to bind tRNA in solution, may be involved in the fixation of the 3' terminus of peptidyl-tRNA at the P-site of the ribosome [39].…”

Section: Discussionmentioning

confidence: 99%

Reconstruction of peptidyltransferase activity on 50S and 70S ribosomal particles by peptide fragments of protein L16

et al. 1987

View full text Add to dashboard Cite

Ribosomal protein L16 was digested with Staphylococcus aureus protease V8 and the resulting peptides were separated by reversed-phase high-performance liquid chromatography. One of the fragments, identified by sequence analysis as the N-terminal peptide of L16, was shown to exhibit partial peptide-bond-formation and transesterification activities of peptidyltransferase upon reconstitution with L16-depleted 50 S core particles. However, several proteins enhanced these activities. L15 increased both reactions when added to the reconstitution mixture, suggesting a limited capacity of the L16 peptide to incorporate into 50s core particles. In contrast, the interaction of L11 with the N-terminal peptide stimulated the transesterification reaction but not the peptidebond-forming activity of ribosomes, indicating a different topological domain for these reactions. Also, EF-P, a soluble protein which reconstructs the peptide-bond formation and transesterification reactions on 70 S ribosomes, stimulated both peptidyltransferase activities exhibited by the L16 N-terminal peptide.Central to the study of the mechanism of peptide-bond synthesis is establishing whether the ribosomal proteins function passively by ensuring correct steric alignment of substrates, or whether they are actively involved in the catalytic process. Thermodynamic considerations and kinetic studies on model compounds with appropriately juxtaposed functional groups indicate that an exclusively template role for the ribosome is at least plausible and may also be sufficient to account for the rate of protein synthesis in vivo [l]. However, several lines of evidence suggest that the ribosomal peptidyltransferase possesses catalytic residue(s) as well [l, 21. Of the known ribosomal activities, ester aminolysis and ester hydrolysis are most likely to be mediated by some form of catalytic mechanism. In this regard, we have recently reported that ribosomal protein L16 stimulates both the peptide-bond formation and transesterification activities inherent in peptidyltransferase [3]. This study confirmed and extended the findings of previous reconstitution experiments which showed that L16 belongs to a group of 50s subunit components designated as essential to restoring the peptidyltransferase activity of ribosomes [4]. In addition, our previous chemical modification studies have pointed to the involvement of an active histidine residue during peptide bond synthesis [5 ~ 71. Although other ribosomal proteins (e.g. L2 and L4) also contain such potentially active residues [2], the unique histidine of L16 has to date remained a prime candidate for providing, at least in part, the catalytic activity of the peptidebond formation and transesterification processes [8]. In this connection, we report here on the reconstruction of both peptidyltransferase activities of 50s and 70s ribosomes by proteolytic fragments of L16 and study the effect of L6, L11 and L15 and the stimulatory protein EF-P on these reactions. MATERIALS AND METHODS MaterialsPuromycin dihydrochloride ...

show abstract

Section: Discussionmentioning

confidence: 99%

Reconstruction of peptidyltransferase activity on 50S and 70S ribosomal particles by peptide fragments of protein L16

et al. 1987

View full text Add to dashboard Cite

show abstract

“…L15 can now be excluded from the candidates for a peptidyltransferase 'enzyme' since an E. coli mutant has been found lacking L15 [22]. L2 has been shown to interact with Met-tRNA and AcPhe-tRNA and to enhance the rate of the spontaneous hydrolysis of the aminoacyl moieties from these tRNAs in solution but not other aminoacyl-tRNAs [23]. L2 is only partially removed from 50s subunits by 1.5 M LiCl and a significant proportion of the 1 .…”

Section: Does the Modified L16 Influence The Assembl-v Of The Peptidymentioning

confidence: 99%

The peptidyltransferase centre of the Escherichia coli ribosome

Tate

Sumpter

Trotman

et al. 1987

European Journal of Biochemistry

View full text Add to dashboard Cite

1. Modification of the Escherichia coli 50s ribosomal subunit with histidine-specific diethyl pyrocarbonate affects peptide bond formation and release-factor-dependent peptidyl-tRNA hydrolysis.2. Unmodified L16 can restore activity to a split protein fraction from the altered subunit but other proteins of the core also contain histidine residues important for the activity of the peptidyltransferase centre, 3. When isolated and purified by centrifugation, particles reconstituted with unmodified proteins and modified L16 do not retain the altered L16. The modified protein does mediate the partial restoration of peptide bond formation and release-factor-2 activities to these particles. It must be exerting its effect during the assembly of the peptidyltransferase centre in the reconstituted particle.4. A particle could be reconstituted which lacks L16 and has significant activity in peptide bond formation and peptidyl-tRNA hydrolysis. L16 stimulates these activities.5. A tighter ribosomal binding of the release factor 2, dependent upon the absence of protein L11, can in part compensate for the loss of activity of the peptidyltransferase centre when it is assembled with either modified L16 or in the absence of L16. The protein and its histidine residue seem important, therefore, for the peptidyltransferase centre to be formed in the correct conformation but not essential for activity once the centre is assembled.The functional and structural integrity of the peptidyltransferase centre of the ribosome is likely to depend on several components including both RNA and proteins. A group of five proteins including L16 is essential for reconstitution of peptide-bond-forming activity to ribosomal core particles [l]. The peptidyl-tRNA hydrolysis step of peptide chain termination, mediated by release factor, is also catalysed by the peptidyltransferase activity of the ribosome [2]. Attempts to define a single component as the enzyme of the centre have been unsuccessful. One model to explain the molecular mechanism of both peptide bond formation and peptidyl-tRNA hydrolysis, however, postulates the participation of histidine and a carboxyl function from a ribosomal component analogous to the active centre of a serine protease [3]. L16 has been a prime candidate for providing at least part of the catalytic function of peptidyltransferase. Indeed modification of the histidyl residue on L16 [4-61 and also those of L2 [7] inactivates the centre. Protein L16 or a proteolytic fragment lacking nine amino acids at the N terminus can restore activity to ribosomal subparticles whereas a fragment lacking a further six amino acids, including the single histidine residue, does not [8, 91. When L16 is missing from 50s reconstituted subunits the particle is inactive for releasefactor-mediated peptidyl-tRNA hydrolysis [lo]. Abbreviations. RF, release factor; 1 . 5~ core, ribosomal particle remaining after extraction of 50s subunits with 1.5 M LiCl; DEP, diethyl pyrocarbonate.Enzyme. Peptidyltransferase (EC 2.3.2.12).While these observations ar...

show abstract

Protein Kinase Associated with Ribosomes Phosphorylates Ribosomal Proteins ofStreptomyces collinus

Mikulı́k

Janda

1997

Biochemical and Biophysical Research Communications

View full text Add to dashboard Cite

The properties of the complex between ribosomal protein L2 and tRNA

Cited by 8 publications

References 17 publications

Reconstruction of peptidyltransferase activity on 50S and 70S ribosomal particles by peptide fragments of protein L16

Reconstruction of peptidyltransferase activity on 50S and 70S ribosomal particles by peptide fragments of protein L16

The peptidyltransferase centre of the Escherichia coli ribosome

Protein Kinase Associated with Ribosomes Phosphorylates Ribosomal Proteins ofStreptomyces collinus

Contact Info

Product

Resources

About