Isolation of tyrosinase-mRNA by affinity chromatography of polysomes.
We present a method whereby some mRNAs which code for enzymes can be isolated by affinity chromatography of newly synthesized polypeptides bound to their polysome complexes. Using this method we have isolated tyrosinase-mRNA from Xenopus laevis oocytes and have analysed the translation products from the RNAs thus obtained. In vitro translation reveals the presence of two mRNAs coding for polypeptides of molecular weights of 20 000 and 32 000, respectively. The larger molecule corresponds to the molecular weight of nascent tyrosinase. Furthermore, microinjection of the mRNA into Xenopus oocytes results in the synthesis of active tyrosinase. Since this isolation method is dependent on the ability of nascent enzymes to bind to their substrate analogue, it is thought that this approach may be appropriate for obtaining mRNAs coding for other enzymes.
D000818:Animals; D004156:Catechol Oxidase / Q000235:genetics*; D002846:Chromatography, Affinity / Q000379:methods*; D005260:Female; D008845:Microinjections; D008970:Molecular Weight; D014442:Monophenol Monooxygenase / Q000235:genetics*; D009865:Oocytes / Q000032:analysis; D014176:Protein Biosynthesis; D012333:RNA, Messenger / Q000302:isolation & purification*; D012270:Ribosomes / Q000032:analysis*; D014982:Xenopus laevis
D012333:RNA, Messenger; D004156:Catechol Oxidase; D014442:Monophenol Monooxygenase