Macromolecular Complexes of Aminoacyl‐tRNA Synthetases from Eukaryotes

Kellermann, Odile; Brevet, Annie; Tonetti, Hubert; Waller, Jean‐Pierre

doi:10.1111/j.1432-1033.1979.tb13286.x

Cited by 70 publications

(42 citation statements)

References 14 publications

(2 reference statements)

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Aminoacyl-tRNA-synthetases were assayed by the aminoacylation of tRNA, as previously described [5]. Unfractionated brewer's yeast tRNA (Boehringer) was used as substrate, except in the case of the aminoacyl-tRNA synthetases specific for arginine, glutamic acid, glutamine, aspartic acid, alanine, asparagine, cysteine, threonine and tryptophan, which were assayed in the presence of partially purified beef liver tRNA [5].…”

Section: Methodsmentioning

confidence: 99%

“…The objectives of the present study were to determine if multisynthetase complexes, similar to those previously purified from sheep and rabbit [5], could be isolated from CHO cells and, if so, to examine the consequences of altered leucyltRNA synthetase on the structure of such complexes in the mutant tsHl cell line. The results presented show that nine aminoacyl-tRNA synthetases, including leucyl-tRNA synthetase, could be copurified from wild-type as well as tsH1 cells to yield a multienzyme complex, the composition of which was identical to that previously determined for the purified complex from rabbit liver.…”

mentioning

confidence: 99%

See 1 more Smart Citation

A complex from cultured Chinese hamster ovary cells containing nine aminoacyl‐tRNA synthetases

Mirande

Corre

Waller

1985

European Journal of Biochemistry

View full text Add to dashboard Cite

The size distribution of the 20 aminoacyl-tRNA synthetases from wild-type Chinese hamster ovary (CHO) cells and from the mutant cell line tsHl , containing a temperature-sensitive leucyl-tRNA synthetase, was determined by gel filtration. Nine aminoacyl-tRNA synthetases, specific for arginine, aspartic acid, glutamic acid, glutamine, isoleucine, leucine, lysine, methionine and proline, which coeluted as high-MI entities ( M r z 1.2 x lo6), were further co-purified to yield a multienzyme complex, the polypeptide composition of which was identical to that previously determined for the complex from rabbit liver. Immunoprecipitates obtained from crude extracts of wild-type and tsHl mutant cells, using specific antibodies directed to the lysyl-tRNA or methionyl-tRNA synthetase components of the complex, displayed the same polypeptide compositions as that of the purified complex, thereby establishing the heterotypic nature of this complex. Although the activity of leucyl-tRNA synthetase from the mutant cells, grown at a permissive temperature, was low compared to that from the wildtype, the polypeptide of M , 129000, corresponding to this enzyme, was present in similar amounts and occurred exclusively as a component of the high-M, complex. Finally, we report that attempts to demonstrate phosphorylation of the components of the complex from cultured CHO, HeLa and C3 cells were unsuccessful.

show abstract

Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

A complex from cultured Chinese hamster ovary cells containing nine aminoacyl‐tRNA synthetases

Mirande

Corre

Waller

1985

European Journal of Biochemistry

View full text Add to dashboard Cite

show abstract

“…The purification procedure was similar to that of Kellermann et al [12]. All steps of the purification procedure were carried out at 4 mC.…”

Section: Purification Of the Multi-enzyme Complex Of Aminoacyl-trna Smentioning

confidence: 99%

“…The final purification step was achieved using affinity chromatography according to the method of Kellermann et al [12]. A Sepharose affinity column containing covalently bound tRNA was equilibrated with buffer B. Fractions containing synthetase activity were pooled and passed twice through this column at a flow rate of 20 ml per h. The column was then washed with 5 ml of buffer B.…”

Section: Purification Of the Multi-enzyme Complex Of Aminoacyl-trna Smentioning

confidence: 99%

See 1 more Smart Citation

A novel pathway for the conversion of homocysteine to methionine in eukaryotes

Antonio

Nunes

Refsum

et al. 1997

Biochemical Journal

View full text Add to dashboard Cite

Activation of amino acid homocysteine was compared with that of methionine in rabbit crude liver extracts and purified multi-enzyme complex of aminoacyl-tRNA synthetases. Activation was studied by measuring the incorporation of radioactive amino acid into unlabelled trichloroacetic-acid insoluble materials in the absence of protein synthesis. Homocysteine synthetase activity was found in the crude extract and in the purified multi-enzyme complex of aminoacyl-tRNA synthetases. On a molar basis, the activation of methionine by the crude extract was five times higher than the activation of homocysteine. There was a partial loss of Hcy-tRNA synthetase activity in the purified multi-enzyme complex. Preliminary reconstitution experiments indicated a requirement for an additional factor for Hcy-tRNA synthetase activity. TLC of the amino acid released from tRNA charged with [14C]homocysteine, revealed radioactivity in homocysteine, methionine and homocysteine thiolactone, indicating a conversion of tRNA-attached homocysteine to methionine. Total tRNA was separated on a benzoylated cellulose column into a fraction enriched in initiator tRNA and a methionine-accepting, but initiator tRNA-deficient, fraction. Homocysteine-accepting activity was present only in the initiator tRNA-enriched fraction. Based on the above data we propose that homocysteine activation in reticulocyte lysates, reported previously, also occurs in liver. Activated homocysteine is attached to initiator tRNA and then converted to methionine by a methylating enzyme. In the absence of methylation, tRNA-attached homocysteine is hydrolysed to produce homocysteine thiolactone.

show abstract

Seven mammalian aminoacyl-tRNA synthetases co-purified as high molecular weight entities are associated within the same complex.

Mirande¹,

Gache²,

Corre³

et al. 1982

The EMBO Journal

View full text Add to dashboard Cite

Seven aminoacyl‐tRNA synthetases from sheep liver were co‐purified as high mol. wt. entities to constant specific activities. The purified multienzyme preparation displayed an apparent mol. wt. of approximately 10(6) and was composed of 11 distinct polypeptides, as revealed by SDS‐polyacrylamide gel electrophoresis (SDS‐PAGE). To test the assumption that all of these components were physically associated within the same complex, the purified preparation was subjected to immunoprecipitation by antibodies raised against its lysyl‐ or methionyl‐tRNA synthetase component. Depending on the limiting concentrations of the specific antibodies used, from 5 to 40% of the input protein was recovered in the immunoprecipitate. Its polypeptide composition, as revealed by SDS‐PAGE, was indistinguishable from that of the original material. The immunoprecipitation reaction was highly specific, as attested by the observation that IgG from nonimmunized rabbit failed to precipitate any of the 11 polypeptides, even when used in 30‐fold molar excess over input protein. We conclude that co‐precipitation of all of these polypeptides by antibodies directed against a single component of the purified preparation is a consequence of their physical association within the same multienzyme complex.

show abstract

Macromolecular Complexes of Aminoacyl‐tRNA Synthetases from Eukaryotes

Cited by 70 publications

References 14 publications

A complex from cultured Chinese hamster ovary cells containing nine aminoacyl‐tRNA synthetases

A complex from cultured Chinese hamster ovary cells containing nine aminoacyl‐tRNA synthetases

A novel pathway for the conversion of homocysteine to methionine in eukaryotes

Seven mammalian aminoacyl-tRNA synthetases co-purified as high molecular weight entities are associated within the same complex.

Contact Info

Product

Resources

About