Mitochondrial peptide chain elongation factors from <i>Neurospora crassa</i>

Grandi, Maria; Küntzel, Hans

doi:10.1016/0014-5793(70)80407-0

Cited by 31 publications

(8 citation statements)

References 10 publications

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“…Previously, only a partial characterization has been reported in the case of the elongation factors present in mitochondria from yeasts [20,21] and Neuvospora CYUSSU [22] while no data at all are available for those present in chloroplasts. We hope that in a not too distant future it will be possible to compare at the molecular level the elongation factors present in the cytoplasm of eukaryotes, in their organelles and in prokaryotes.…”

Section: Discussionmentioning

confidence: 99%

Purification of the Elongation Factors Present in Spinach Chloroplasts

Tiboni

Pasquale

Ciferri

1978

European Journal of Biochemistry

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Elongation factor G (EF-Gchl) and elongation factor Tu (EF-Tuchl) have been purified from isolated spinach chloroplasts. On polyacrylamide gel electrophoresis the purified proteins appear to be at least 70% pure. The molecular weight has been estimated to be 77000 and 45500 for EF-Gchl and EF-Tuchl respectively.Chloroplast elongation factor T (EF-Tchl) has been only partially purified. Gel electrophoresis under non-denaturing and denaturing conditions indicate that EF-Tchl is most probably composed of two polypeptides, one of which has an electrophoretic mobility identical to that of EF-Tuchl.EF-Tuchl appears to represent approximately 7 % of the chloroplast soluble protein while EF-Gchl accounts for less than 1 %. Just as in the case of the bacterial factors, EF-Tuchl appears to be in excess as compared to EF-Gchl.Although no data were obtained on the concentration of EF-Tchl, it may be assumed that the three elongation factors represent approximately 10 % of the chloroplast soluble protein.It is well known that chloroplasts synthesize protein and that the protein-synthetic apparatus of these organelles is different from that present in the cell cytoplasm [1,2]. Perhaps the most striking difference is the 'prokaryotic' type of the machinery for protein synthesis in chloroplasts as compared to the 'eukaryotic' type of that present in the cell cytoplasm. Indeed, just like prokaryotes, chloroplasts contain ribosomes of the 70-S type, peptide chains initiate in these organelles with formylated methionine and protein synthesis is inhibited by antibiotics such as chloramphenicol that inhibit this process in prokaryotes but not by typical inhibitors of protein synthesis in the cytoplasm of eukaryotes such as cycloheximide In previous papers we have shown that chloroplasts contain specific elongation factors differing from those present in the cell cytoplasm and in mitochondria [5-71. In this paper we report the purification of the elongation factors present in spinach chloroplasts. To our knowledge this is the first report of the purification and the characterization of these factors from chloroplasts.

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Section: Discussionmentioning

confidence: 99%

Purification of the Elongation Factors Present in Spinach Chloroplasts

Tiboni

Pasquale

Ciferri

1978

European Journal of Biochemistry

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“…In the bacterial system mitochondrial peptide chain elongation factors from yeast and Neurosporu were successfully substituted in place of Escherichiu coli factors [6,7], but the reverse experiment appeared to be limited by the low activity of isolated mitochondrial ribosomes in protein synthesis in vitro. Isolation of active mitochondrial ribosomes free of endogenous elongation factors therefore was a prerequisite to study the specificity of bacterial and mitochondrial peptide chain elongation factors.…”

Section: Mammalian Mitochondrial Ribosomes Studies On the Exchangeabimentioning

confidence: 99%

“…Although differences have been observed between ribosomes from bacteria and mammalian mitochondria in the interaction with antibiotics binding to the ribosomal P-site, 55-S ribosomes can be classed as procaryotic by functional parameters [5].The degree of similarity between bacterial and mitochondrial mechanisms of protein synthesis can be expressed by the interchangeability of protein synthesis factors, ribosomal proteins, or even ribosomal subunits. In the bacterial system mitochondrial peptide chain elongation factors from yeast and Neurosporu were successfully substituted in place of Escherichiu coli factors [6,7], but the reverse experiment appeared to be limited by the low activity of isolated mitochondrial ribosomes in protein synthesis in vitro.Abbreviation. Tes, 2-( [2-hydroxy-I ,I -bis(hydroxymethyl)ethyl]-amino}ethanesulfonic acid.Isolation of active mitochondrial ribosomes free of endogenous elongation factors therefore was a prerequisite to study the specificity of bacterial and mitochondrial peptide chain elongation factors.…”

mentioning

confidence: 99%

“…The degree of similarity between bacterial and mitochondrial mechanisms of protein synthesis can be expressed by the interchangeability of protein synthesis factors, ribosomal proteins, or even ribosomal subunits. In the bacterial system mitochondrial peptide chain elongation factors from yeast and Neurosporu were successfully substituted in place of Escherichiu coli factors [6,7], but the reverse experiment appeared to be limited by the low activity of isolated mitochondrial ribosomes in protein synthesis in vitro.…”

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confidence: 99%

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Mammalian Mitochondrial Ribosomes. Studies on the Exchangeability of Polypeptide Chain Elongation Factors from Bacterial and Mitochondrial Systems

1980

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Mammalian mitochondrial ribosomes from rat liver synthesised poly(phenyla1anine) from ['"CIPhe-tRNA in the presence of a homologous lo5 x g,, supernatent fraction. The activity depended on the addition of synthetic template and was resistant to cycloheximide. The polyanion spermidine had a stimulatory effect on peptide synthesis in vitro. In contrast to Escherichiu coli ribosomes, which also functioned with heterologous supernatant fractions, 55-S mitochondrial ribosomes were inactive when supplemented with heterologous supernatant fractions from E. coli or with purified bacterial elongation factors. EF-T slightly stimulated polyphenylalanine synthesis when added in combination with mitochondrial supernatant fractions. Two-dimensional electrophoretic analysis of the protein content of both supernatant fractions revealed considerable differences in the distribution of the species-specific proteins according to their isoelectric points. The mitochondrial superand the few acidic proteins did not co-migrate with natant proteins were in general more basic EF-Tu or EF-G from E. coli.Mammalian mitochondrial ribosomes represent a special group among organelle ribosomes. They display an unusually high protein content [l -31 and evolved fairly rapidly compared with cytoplasmic ribosomes [4]. Like all other mitochondrial ribosomes they contain binding sites for inhibitors of bacterial protein synthesis while being unaffected by antibiotics such as cycloheximide and anisomycin which interfere with eucaryotic ribosome function. Although differences have been observed between ribosomes from bacteria and mammalian mitochondria in the interaction with antibiotics binding to the ribosomal P-site, 55-S ribosomes can be classed as procaryotic by functional parameters [5].The degree of similarity between bacterial and mitochondrial mechanisms of protein synthesis can be expressed by the interchangeability of protein synthesis factors, ribosomal proteins, or even ribosomal subunits. In the bacterial system mitochondrial peptide chain elongation factors from yeast and Neurosporu were successfully substituted in place of Escherichiu coli factors [6,7], but the reverse experiment appeared to be limited by the low activity of isolated mitochondrial ribosomes in protein synthesis in vitro. Isolation of active mitochondrial ribosomes free of endogenous elongation factors therefore was a prerequisite to study the specificity of bacterial and mitochondrial peptide chain elongation factors. The present investigation reports on the ability of supernatant fractions from rat liver mitochondria and from E. coli to stimulate peptide synthesis in homologous and heterologous systems. The protein composition of both 1 O5 x g,, supernatant fractions was compared by two-dimensional gel electrophoresis applying isoelectric focusing in the first dimension. MATERIALS AND METHODS MateriulsRadioactive amino acids and [3H]poly(U) were obtained from Amersham, Great Britain. A11 reagents were of analytical grade. Solutions were prepared in sterilized glasswar...

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“…Use of methods for the preparation of bacterial factors has enabled the purification of T and G factors from yeast and Neurospora mitochondria to be accomplished, and the ability of these factors to support phenylalanine polymerization on E. coli ribosomes in combination with the complementary E. coli factors, has been investigated (Grandi & Kuntzel, 1970; Morimoto et al 1971). The combination of T (E. coli) and G (Neurospora) was found to be active (Grandi & Kuntzel, 1970), as was that of T, (yeast) and G (E. coli) (Morimoto et al 1 9 7 1 ) ; the lability of mitochondrial T factors was probably responsible for T (Neurospora) and G (E. coli) not displaying any activity (Grandi & Kuntzel, 1970).…”

Section: ( 2 ) Transfer Rnas Aminoacyl-trna Synthetasesmentioning

confidence: 95%