The many applications of acid urea polyacrylamide gel electrophoresis to studies of tRNAs and aminoacyl-tRNA synthetases.
Here we describe the many applications of acid urea polyacrylamide gel electrophoresis (acid urea PAGE) followed by Northern blot analysis to studies of tRNAs and aminoacyl-tRNA synthetases. Acid urea PAGE allows the electrophoretic separation of different forms of a tRNA, discriminated by changes in bulk, charge, and/or conformation that are brought about by aminoacylation, formylation, or modification of a tRNA. Among the examples described are (i) analysis of the effect of mutations in the Escherichia coli initiator tRNA on its aminoacylation and formylation; (ii) evidence of orthogonality of suppressor tRNAs in mammalian cells and yeast; (iii) analysis of aminoacylation specificity of an archaeal prolyl-tRNA synthetase that can aminoacylate archaeal tRNA(Pro) with cysteine, but does not aminoacylate archaeal tRNA(Cys) with cysteine; (iv) identification and characterization of the AUA-decoding minor tRNA(Ile) in archaea; and (v) evidence that the archaeal minor tRNA(Ile) contains a modified base in the wobble position different from lysidine found in the corresponding eubacterial tRNA.
Köhrer
Caroline
C
Department of Biology, Massachusetts Institute of Technology, 77 Massachusetts Avenue, Room 68-671, Cambridge, MA 02139, USA.
Rajbhandary
Uttam L
UL
D000604:Amino Acyl-tRNA Synthetases / Q000032:analysis*; D000818:Animals; D001105:Archaea / Q000378:metabolism; D015152:Blotting, Northern / Q000379:methods; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008239:Lysine / Q000031:analogs & derivatives / Q000096:biosynthesis; D015202:Protein Engineering / Q000379:methods; D011741:Pyrimidine Nucleosides / Q000096:biosynthesis; D012329:RNA, Bacterial / Q000302:isolation & purification; D012343:RNA, Transfer / Q000032:analysis* / Q000302:isolation & purification; D012350:RNA, Transfer, Cys / Q000096:biosynthesis; D012355:RNA, Transfer, Ile / Q000378:metabolism; D012358:RNA, Transfer, Met / Q000378:metabolism; D014508:Urea
D011741:Pyrimidine Nucleosides; D012329:RNA, Bacterial; D012350:RNA, Transfer, Cys; D012355:RNA, Transfer, Ile; D012358:RNA, Transfer, Met; C024988:tRNA, formylmethionine-; D014508:Urea; D012343:RNA, Transfer; C101421:lysidine; D000604:Amino Acyl-tRNA Synthetases; D008239:Lysine