Rapid purification of tRNA(Lys) from rat liver.
Biochem Mol Biol Int
; 33(6): 1081-9, 1994 Aug.
Article
em En
| MEDLINE
| ID: mdl-7804133
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.
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Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
RNA de Transferência de Lisina
/
Fígado
Limite:
Animals
Idioma:
En
Revista:
Biochem Mol Biol Int
Assunto da revista:
BIOLOGIA MOLECULAR
/
BIOQUIMICA
Ano de publicação:
1994
Tipo de documento:
Article