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Rapid purification of tRNA(Lys) from rat liver

Kumar, AM and Vulimiri, SV and Nayak, R (1994) Rapid purification of tRNA(Lys) from rat liver. In: Biochemistry & Molecular Biology International, 33 (6). pp. 1081-1089.

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Abstract

Fast protein liquid chromatography (FPLC) system using Mono Q (HR 5/5) anion-exchange column chromatography followed by highly cross-linked urea-polyacrylamide gel electrophoresis (urea-PAGE) was used for the purification of lysine-specific tRNA (tRNA(Lys)) from rat liver. Crude tRNA from rat liver was fractionated with a linear gradient of NaCl (0.3-0.8 M) in triethanolamine-HCl buffer, pH 4.5, and the activity of tRNA(Lys) was found to elute between 0.51 and 0.57 M NaCl. Using this concentration range of NaCl, tRNA(Lys) was refractionated on the same column with a shallow gradient, where a single peak of tRNA(Lys) activity was obtained. tRNA(Lys)-rich fractions recovered from the second run were electrophoretically separated on 16% polyacrylamide-7 M urea gel into one major band and three minor bands. The major band showed a specific activity of 997 pmols/A260 U for tRNALys with a 43-fold purification and approximately 17% recovery. The minor bands displayed negligible or no activity for lysine. tRNA(Lys) obtained by this method was found to be homogeneous by competitive aminoacylation. The advantages of FPLC followed by urea-PAGE in the purification of an amino acid-specific tRNA over conventional column chromatography are discussed.

Item Type: Journal Article
Additional Information: Copyright of this article belongs to Academic press aust.
Department/Centre: Division of Biological Sciences > Microbiology & Cell Biology
Date Deposited: 11 Apr 2011 10:34
Last Modified: 11 Apr 2011 10:34
URI: http://eprints.iisc.ernet.in/id/eprint/36717

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