Further Evidence that Elongation Factor 1 Remains Bound to Ribosomes during Peptide Chain Elongation

Grasmuk, Hans; Nolan, Robert D.; Drews, Jürgen

doi:10.1111/j.1432-1033.1977.tb11787.x

Cited by 24 publications

(13 citation statements)

References 34 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Using diphtheria toxin-mediated ADP-ribosylation to assay for EF-2, it was shown that inhibition of EF-2 binding to pre-translocative ribosome Á EF-1a complexes prevented ribosomal translocation. Employing didemnin B as a tool, evidence was provided that EF-1a remains bound to the ribosome throughout the elongation cycle consistent with the model proposed previously by Grasmuk et al 123,124 and supported by other authors. 125 In this model, two mechanisms for inhibition of EF-2 binding may be considered: a competitive mechanism in which didemnin B or EF-1a occupy the ribosome where EF-2 normally binds, or an uncompetitive mechanism in which didemnin B binds to ribosome Á EF-1a complex and locks the complex in a conformation that disfavors EF-2 binding.…”

Section: Didemnin Binding Proteins Ef-1a and Ppt1supporting

confidence: 70%

Natural products as probes of cell biology: 20 years of didemnin research

Vera

Joullié

2002

Medicinal Research Reviews

143

109

View full text Add to dashboard Cite

The discovery of the didemnin family of marine depsipeptides launched an exciting and intriguing chapter in natural product chemistry. The unusual structure of the didemnin congeners has led to several total syntheses by research groups from around the world. The impressive in vitro and in vivo biological activities of the didemnins resulted in the first human clinical trials in the U.S. of a marine natural product against cancer, and additional clinical trials of a second-generation didemnin, dehydrodidemnin B (aplidine), are underway. As we mark the 20-year anniversary of the discovery of the didemnins, this class of natural products continues to stimulate active research in fields ranging from synthetic and medicinal chemistry to clinical oncology and cell biology. While some progress was made in dissecting the molecular mechanism of action and in establishing structure-activity relationships, there are still more questions than answers. This review covers the recent didemnin literature, highlighting the work directed towards understanding how this group of natural products interact with fundamental processes such as cell proliferation, protein biosynthesis, and apoptosis. The didemnin field illustrates how natural product chemistry may be used as a critical tool for the study of cell biology.

show abstract

Section: Didemnin Binding Proteins Ef-1a and Ppt1supporting

confidence: 70%

Natural products as probes of cell biology: 20 years of didemnin research

Vera

Joullié

2002

Medicinal Research Reviews

143

109

View full text Add to dashboard Cite

show abstract

“…The N-terminal sequence of P2 is analogous to that of Escherichia coli L7/L12 [35, 361. Furthermore, employing ribosomes and EF-2 from ascites tumor cells, Gasmuck et al [37] reported that antiserum against E. coli L7/L12 proteins inhibited binding of [3H]EF-2 Table 1. Summary ofproteins cross-linked to EF-2 with 2-iminothiolane The relative molecular masses of cross-linked protein dimers of EF-2 -ribosomal-protein were calculated from their migrations in the first dimension on diagonal gel electrophoresis.…”

mentioning

confidence: 99%

mentioning

confidence: 99%

Cross-linking of elongation factor 2 to rat-liver ribosomal proteins by 2-iminothiolane

et al. 1986

View full text Add to dashboard Cite

Complexes containing rat liver 80s ribosomes treated with puromycin and high concentrations of KCl, elongation factor 2 (EF-2) from pig liver, and guanosine 5'-[/3,y-methylene]triphosphate were prepared. Neighboring proteins in the complexes were cross-linked with the bifunctional reagent 2-iminothiolane. Proteins were extracted and then separated into 22 fractions by chromatography on carboxymethylcellulose of which seven fractions were used for further analyses. Each protein fraction was subjected to diagonal polyacrylamide/sodium dodecyl sulfate gel electrophoresis. Nine cross-linked protein pairs between EF-2 and ribosomal proteins were shifted from the line formed with monomeric proteins. The spots of ribosomal proteins cross-linked to EF-2 were cut out from the gel plate and labelled with lZ5I. The labelled protein was extracted from the gel and identified by three kinds of two-dimensional gel electrophoresis, followed by autoradiography. The following proteins of both large and small subunits were identified: L9, L12, L23, LA33 (acidic protein of M, 33000), P2, S6 and S23/ S24, and L3 and L4 in lower yields. The results are discussed in relation to the topographies of ribosomal proteins in large and small subunits. Furthermore we found new neighboring protein pairs in large subunits, LA33 -L11 and LA33 -L12.During protein biosynthesis in eukaryotic cells, elongation factor 2 (EF-2) interacts with ribosomes in the presence of GTP and catalyzes the translocation of newly synthesized peptidyl-tRNA from the aminoacyl-tRNA binding site (the A site) of ribosomes to the peptidyl-tRNA site (the P site) [l -31. The functional roles of ribosomal proteins in the translocation are as yet unknown. To understand the molecular mechanisms of the translocation in mammalian ribosomes, it is important to identify proteins in the EF-2 binding site of ribosomes.Employing four kinds of cross-linking reagents and hydrogen peroxide, we identified 62 cross-linked protein pairs involving 36 protein species containing two acidic proteins of rat liver 60s subunits and have discussed these in relation to other functional data [4, 51. Concerning the 40s subunits, similar data were presented by Tolan and Traut [6]. Therefore, it is of interest that ribosomal proteins which interact with elongation factors have been determined.In the present experiments we investigated EF-2-binding sites in rat liver 80s ribosomes, employing 2-iminothiolane. Cross-linked proteins were analyzed by a modified method of diagonal SDS gel electrophoresis [7]. Seven large-subunit proteins and two small-subunit proteins were identified as the components cross-linked to EF-2. The mutual locations of these proteins in the 80s ribosomes are discussed.

show abstract

“…These initial reports came from studies on the silk worm [4], pig liver [ 5 ] , and more recently on systems derived from the cysts of the brine shrimp, Artemia salina [l]. Intensive efforts in the past to observe such an activity in the Krebs I1 mouse ascites tumor cell system failed although eEF-Ts from Arternia showed stimulating activity in the ascites cell-free protein-synthesizing system [6]. This evidence led us to the conclusion that the activity was not to be found in ascites cells.…”

mentioning

confidence: 99%

The Isolation and Characterization of Elongation Factor eEF‐Ts from Krebs‐II Mouse‐Ascites‐Tumor Cells and Its Role in the Elongation Process

Grasmuk

Nolan

Drews

1978

European Journal of Biochemistry

Self Cite

View full text Add to dashboard Cite

A factor having activity similar to that described in other systems for the eukaryotic elongation factor eEF‐Ts was isolated from the heavy, aggregate form of eEF‐TH (formally named EF‐1H). This protein has a molecular weight of 52000 under native conditions and of 25 500 under denaturing conditions. It has been shown to stimulate eEF‐Tu‐dependent aminoacyl‐tRNA binding to ribosomes and therefore eEF‐Tu/eEF‐G‐dependent polyphenylalanine synthesis by ribosomes and was found to stimulate GDP‐GTP exchange in eEF‐Tu · GDP complexes. In the course of this work, it was also demonstrated that the removal of deacylated tRNA from the ribosome is a GTP‐dependent process. This report, therefore, adds further support to the concept that a third elongation factor, eEF‐Ts, may be common to all systems in the eukaryotic domain.

show abstract

Further Evidence that Elongation Factor 1 Remains Bound to Ribosomes during Peptide Chain Elongation

Cited by 24 publications

References 34 publications

Natural products as probes of cell biology: 20 years of didemnin research

Natural products as probes of cell biology: 20 years of didemnin research

Cross-linking of elongation factor 2 to rat-liver ribosomal proteins by 2-iminothiolane

The Isolation and Characterization of Elongation Factor eEF‐Ts from Krebs‐II Mouse‐Ascites‐Tumor Cells and Its Role in the Elongation Process

Contact Info

Product

Resources

About