Studies on the structure and function of E. coli phenylalanine transfer RNA

Lowdon, Margaret (1976) Studies on the structure and function of E. coli phenylalanine transfer RNA. PhD thesis, University of Glasgow.

Abstract

Two different methods have been used for the purification of E. coli TRNA[Phe2]. Both methods involved initial fractionation of unpurified E. coli tRNA (either unlabelled, or uniformly labelled with [32]P) on Benzoylated DEAE-cellulose. The fractions containing TRNA[Phe2] (the second phenylalanine isoacceptor tRNA eluted from the Benzoylated DEAE-cellulose column on increasing the NaC1 concentration of the eluate) were then fractionated either after charging of the tRNA with phenylalanine on another Benzoylated DEAE-cellulose column, or Rhe on an RPC-5 column. After deacylation of purified Phe-tRNA[Phe2] the tRNA[Phe2] could not be charged with phenylalanine to the original extent. Purified tRNA[Phe2] obtained from an RPC-5 column was found to lose phenylalanine accepting activity on storage. Attempts were made to renature both inactivated forms of tRNA[Phe2] but they were not successful. Attempts to fully charge the inactivated tRNA[Phe2] by increasing the ligase concentration in the assay or by preincubating the tRNA[Phe2] with the ligase were equally unsuccessful. However, after investigation of the stability of tRNA[Phe2] under various conditions, it was found to be most stable in 10mr1 MgC12, 10mM tris-HC1, pH 7.0, in the presence of an equal amount of unpurified E. coli tRNA. The purified E. coli tRNA[Phe2] isolated was found to give identical T1 and Pancreatic RNase fingerprints to the E. coli tRNA[Phe] whose sequence was elucidated by Barrel & Sanger (1969). Treatment of E. coli tRNA[Phe2] with sodium bisulphite at 25

• Item Type: Thesis (PhD)
• Qualification Level: Doctoral
• Additional Information: Adviser: J P Goddard
• Keywords: Biochemistry
• Date of Award: 1976
• Depositing User: Enlighten Team
• Unique ID: glathesis:1976-72948
• Copyright: Copyright of this thesis is held by the author.
• Date Deposited: 11 Jun 2019 11:06
• Last Modified: 11 Jun 2019 11:06
• URI: https://theses.gla.ac.uk/id/eprint/72948