The primary structure of non-initiator methionine transfer ribonucleic acid from Bakers' yeast. I. Purification and complete digestion with ribonuclease T1 and pancreatic ribonuclease A.
The methionine acceptor activity of a crude tRNA from bakers' yeast was resolved into two peaks (I and II) by column chromatography on DEAE-Sephadex A-25 with a 1 M phosphate system. Methionine tRNA from peak II was not formylated by E. coli methionyl-tRNA transformylase [EC 2.1.2.9.] after being charged with methionine, whereas that from peak I was formylatable under the same conditions. A substantial amount of unlabelled methionine tRNA, tRNAMetm, was highly purified from the peak II fraction by successive chromatographic procedures. The purified tRNAMetm was digested with pancreatic ribonuclease A [EC 3.1.4.22] and ribonuclease T1 [EC 3.1.4.8]. The digestion products were isolated into individual components and completely sequenced. The results of sequence analysis of the two RNase digests were in good agreement and indicated that the chain length of this tRNA is 76, including 13 modified nucleotides. These oligonucleotide fragments can be constructed into a unique total sequence, assuming a few conventional features of clover leaf structure for the tRNA was established by analyses of partial digestion products with RNase T1, as reported in the accompanying paper.
D001483:Base Sequence; D004926:Escherichia coli / Q000201:enzymology; D005561:Formates / Q000378:metabolism; D008715:Methionine / Q000032:analysis; D009843:Oligoribonucleotides / Q000032:analysis; D012343:RNA, Transfer / Q000032:analysis* / Q000302:isolation & purification; D006163:Ribonuclease T1 / Q000378:metabolism; D012260:Ribonucleases / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000032:analysis*; D014166:Transferases / Q000378:metabolism
D005561:Formates; D009843:Oligoribonucleotides; D012343:RNA, Transfer; D008715:Methionine; D014166:Transferases; D012260:Ribonucleases; D006163:Ribonuclease T1