caa a22 4500 397558589 CHVBK 20180308164810.0 cr unu---uuuuu 161202e199611 xx s 000 0 eng 10.1093/oxfordjournals.jbchem.a021500 doi (NATIONALLICENCE)oxford-10.1093/oxfordjournals.jbchem.a021500 Further Study on Association of 5SrRNA-L5 Protein Complex and Methionyl-tRNA to Methionyl-tRNA Synthetase in the Macromolecular Aminoacyl-tRNA Synthetase Complex [Elektronische Daten] [Kikuo Ogata, Rie Ohno, Sakura Morioka, Kazuo Terao] To obtain direct evidence for the attachment of 5SrRNA-ribosomal L5 protein particles (5SRNP) and methionine-tRNA (tRNAmet) to methionyl-tRNA synthetase (MetRS) in the macromolecular aminoacyl-tRNA synthetase (ARS) complex of rat liver, a MetRS-5SRNP- tRNAmet complex was dissociated from the macromolecular ARS complex fraction by n-octyl-β.D-glucoside (Method I) or by ω-aminooctyl agarose (Method II) chromatography. The dissociated MetRS complex fraction was purified by gel filtration followed by tRNA-Sepharose chromatography using partially purified tRNAmet in Method I, and by hydrophobic interaction chromatography in Method II. In both methods, final SuperdexTM200 chromatography showed that MetRS activity was present in the region corresponding to the molecular weight of the MetRS-5SRNP-tRNAmet complex (Mr 200,000). One main protein band corresponding to the molecular weight of MetRS was observed on SDS-PAGE of the final product, which was concentrated by lyophilizing after dialysis against water. Using serum albumin as an inhibitor of adhesion of L5 to the microconcentrators which was used to concentrate the final product, a distinct L5 band was detected on SDS-PAGE, the intensity of which was comparable to that of the MetRS band. Northern blot analysis of RNA prepared from the tRNA-Sepharose fraction showed the presence of 5SrRNA. Dot blot analysis using an antibody against ribosomal protein L5 showed that L5 was present in the SuperdexTM200 fractions prepared by both methods. The MetRS specific activities in MetRS complex fractions incubated without tRNA increased during the purification procedures, indicating that endogeneous tRNAmet exists stably in the MetRS complex. 5SRNP and 5SrRNA markedly enhanced the MetRS activity in the MetRS complex, indicating that 5SRNP(A) plays a role as a positive effector of MetRS. © 1996 BY THE JOURNAL OF BIOCHEMISTRY Regular Papers nationallicence methionyl-tRNA nationallicence methionyl-tRNA synthetase nationallicence ribosomal protein L5 nationallicence 5S ribosomal RNA nationallicence 5SrRNA-L5 complex nationallicence Ogata Kikuo Institute for Gene Expression, Dobashi Kyoritsu Hospital Dobashi, Matsuyama, Ehime 790 aut Ohno Rie Institute for Gene Expression, Dobashi Kyoritsu Hospital Dobashi, Matsuyama, Ehime 790 aut Morioka Sakura Institute for Gene Expression, Dobashi Kyoritsu Hospital Dobashi, Matsuyama, Ehime 790 aut Terao Kazuo Niigata Women's College, Ebigase Niigata 950 aut The Journal of Biochemistry Oxford University Press 120/5(1996-11), 869-880 0021-924X 120:5<869 1996 120 jbchem https://doi.org/10.1093/oxfordjournals.jbchem.a021500 text/html Onlinezugriff via DOI 1 other jats NATIONALLICENCE 856 40 https://doi.org/10.1093/oxfordjournals.jbchem.a021500 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Ogata Kikuo Institute for Gene Expression, Dobashi Kyoritsu Hospital Dobashi, Matsuyama, Ehime 790 aut NATIONALLICENCE 700 1- Ohno Rie Institute for Gene Expression, Dobashi Kyoritsu Hospital Dobashi, Matsuyama, Ehime 790 aut NATIONALLICENCE 700 1- Morioka Sakura Institute for Gene Expression, Dobashi Kyoritsu Hospital Dobashi, Matsuyama, Ehime 790 aut NATIONALLICENCE 700 1- Terao Kazuo Niigata Women's College, Ebigase Niigata 950 aut NATIONALLICENCE 773 0- The Journal of Biochemistry Oxford University Press 120/5(1996-11), 869-880 0021-924X 120:5<869 1996 120 jbchem Metadata rights reserved CC BY-NC-4.0 http://creativecommons.org/licenses/by-nc/4.0 nationallicence BK010053 XK010053 XK010000 NATIONALLICENCE NATIONALLICENCE NL-oxford