In the Nirenberg and Matthaei experiment the artificial mRNA, polyU, was translated into polyphenylalanine in a cell-free system, establishing that UUU was the codon for Phe. How did this work as the artificial mRNA lacked an AUG initiation codon, which we now know is required for translation of cellular mRNAs?

  • 1
    I have modified your question, eliminating the question of whether there was AUG present. Obviously not — as this was the experiment that established the first codon and nothing was known about AUG and initiation. PolyU was PolyU was PolyU. I think I have clarified your question (as well as answering it below). If not, let me know. – David Jan 22 '17 at 22:33
  • 1
    [1-4]. The story of this experiment is well told by Judson in The Eight Day of Creation (the chapter 'He wasn't a member of the club'). Some quotes: "All through the previous winter, he [Jacob] and [Francois] Gros had attempted to isolate messenger RNA from induced E.coli, put it into a cell-free system , and make enzyme. 'Which we never got', he said. 'But we used to talk as a joke of putting in poly-A or poly-U -' He laughed. 'Oh yes, but is was a joke; I mean, we were absolutely convinced that nothing would have come of that'. – user1136 Jan 23 '17 at 14:26
  • [2-4]. "Tissiers felt a congruent chagrin. 'Of course , the idiotic thing as far as we were concerned is that we had Paul Doty in the lab next door,' he said. 'We were on very good terms; we were talking about science constantly, with Paul Doty. He had poly-U. But we never put is in'. " – user1136 Jan 23 '17 at 14:30
  • [3-4]. "Brenner had not gone to Moscow. 'But I had chaps doing this here, you see,' he said. 'Doing it with turnip-yellow-mosaic virus. You see we said if the message was true, it should be possible to add a message to the ribosomes' But had they not used artificial RNAs? ' No, we didn't use - but I mean that thing, that was done as a control, Brenner said. 'Because we were doing it with tobacco mosaic virus' And poly-U was suggested by Gordon Tomkins as a control. Okay? It didn't occur to us to use synthetic polyers' ". – user1136 Jan 23 '17 at 14:36
  • [4-4] "Severo Ochoa had not gone to Moscow. He had intended to use artificial RNAs as messengers, and was even then refining a cell-free system like Nirenberg's. 'When we heard the news from Moscow, we immediately tried it. And other polymers and co-polymers we had in the icebox' We immediately got results with four or five' ". – user1136 Jan 23 '17 at 14:40

This is an intelligent question that highlights the ‘sleight of hand’ simplifications employed in many text books to make experimental science appear cleaner and more logical than is in fact the case. From what we now know about the requirement for an AUG initiation codon in protein biosynthesis these experiments should not have worked.

The reason they did was that they were performed at an unphysiologically high concentration of Mg2+ which overcomes the need for specific initiation (and, incidentally, allows other reactions of protein synthesis to proceed in an abnormal manner, particularly the codon-directed tRNA binding reaction used to finish the deciphering of the genetic code). You should also bear in mind that the efficiency of the reaction was probably much less than that within the cell, but this was of no concern as all that was required was that a product be synthesised.

Reference

I worked in this area in the late 60s/early 70s and the importance of magnesium concentration was just common knowledge in the field, so it is difficult to cite a reference to support my assertion. However a brief search brought up a contemporay paper on the effect of magnesium on initiation of protein synthesis in phage RNA, illustrating its importance. There are, no doubt, others.

Your Answer

 
discard

By clicking "Post Your Answer", you acknowledge that you have read our updated terms of service, privacy policy and cookie policy, and that your continued use of the website is subject to these policies.

Not the answer you're looking for? Browse other questions tagged or ask your own question.