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TMV: the direction of assembly


Finding the origin of the assembly of TMV
Aaron Klug Scientist
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I remember going to a meeting of the British Biophysical Society in 1971, ahead of me was walking David Phillips and somebody else. And I heard them say, 'Klug is talking about the assembly of TMV', he said, 'What is there to say, it's all understood.' Well, I did speak about this and this was the... I spoke about this and we published some papers in Nature and New Biology, which was the first issue. The journal went defunct, it was the first time Nature had gone into a different field, it was now in many new journals of Nature. So we published these papers on the polymorphic forms of TMV protein and the assembly... the disc aided assembly of TMV protein; so this kind of biological role... and was very pleasing. And it meant, you see it now, the other thing that Joe went on to show is that if you had... if you added... if you try to do the assembly with foreign RNAs not viral RNA, it didn't work. So it was clear that the disc was not only an obligatory intermediate, and Don Caspar had argued there had to be such an obligatory, but, now we had one. But this was also one which distinguished... the RNA of the virus from any other RNA, because if you used yeast RNA or any other RNA or ribosomal RNA, they didn't assemble. And this was, you see, the reason it was important... was because it was believed by a lot of people, because of the paper by Roger Hart who worked with Robley Williams, that TMV protein could assemble with any piece of RNA, there was no specificity. Now... so this explained his observations and this was being echoed by David Baltimore who was trying to assemble polio virus out of protein RNA. And he found he could always encapsulate it under a wide range of conditions, foreign RNAs. So that sort of added to the idea that RNAs, that the... there was no specificity in the interaction.

Well, it turned out it was Rober... Roger Hart had been doing, he'd been doing the assembly at low pH. So what was happening was he was making the... he was making the viral helix, protein helix, there was an array and it was schlurping up, so to speak, the RNA, just sucking it up. And so it was under... but if you worked at pH7 then it was highly specific, so this ment that it was a rather important biological point. It meant the disc was not only involved physically in the assembly, in nucleating the assembly, it was also discriminating between... TMV RNA and foreign RNA, and this is vital for specificity because in the cell there'll be other RNAs and we... and the discs did that. And then later, so how... how did that do that discrimination, well, the thing was to find the origin of assembly, where which of the RNA was being recognised. And this was done by Joe Butler and David Zimmen who joined later; this was done a few years later. They found that the origin of assembly in the RNA was about a thousand bases from the three-prime end... and it was a bit mysterious, this origin of assembly. First of all, it didn't, they... it wasn't the full 6,000 bases of RNA. So although this might be the origin of assembly, the... the, you could only assemble... assembly was going from three to a five-prime direction. It left a mystery about the 1,000 bases around the other end. The... so the... anyway they did, they did the experiment... and they found by stopping the assembly at the early stages, they found that a piece of RNA got encapsulated in the first few seconds or minutes of the assembly. And they found this was a piece about 1,000 bases from the end. And when you wrote out its formula, the RNA formula, you could see that it had formed a hairpin with a loop. And on the loop there was a series of guanines, three bases apart, there were five guanines. Now, there are three amino acids for... sorry three nucleotides for every amino acid in the helical array and this probably meant that the guanine was a signal for the recognition by the... by the proteins. So... And so, that solved that problem.

Born in Lithuania, Aaron Klug (1926-2018) was a British chemist and biophysicist. He was awarded the Nobel Prize in Chemistry in 1982 for developments in electron microscopy and his work on complexes of nucleic acids and proteins. He studied crystallography at the University of Cape Town before moving to England, completing his doctorate in 1953 at Trinity College, Cambridge. In 1981, he was awarded the Louisa Gross Horwitz Prize from Columbia University. His long and influential career led to a knighthood in 1988. He was also elected President of the Royal Society, and served there from 1995-2000.

Listeners: Ken Holmes John Finch

Kenneth Holmes was born in London in 1934 and attended schools in Chiswick. He obtained his BA at St Johns College, Cambridge. He obtained his PhD at Birkbeck College, London working on the structure of tobacco mosaic virus with Rosalind Franklin and Aaron Klug. After a post-doc at Childrens' Hospital, Boston, where he started to work on muscle structure, he joined to the newly opened Laboratory of Molecular Biology in Cambridge where he stayed for six years. He worked with Aaron Klug on virus structure and with Hugh Huxley on muscle. He then moved to Heidelberg to open the Department of Biophysics at the Max Planck Institute for Medical Research where he remained as director until his retirement. During this time he completed the structure of tobacco mosaic virus and solved the structures of a number of protein molecules including the structure of the muscle protein actin and the actin filament. Recently he has worked on the molecular mechanism of muscle contraction. He also initiated the use of synchrotron radiation as a source for X-ray diffraction and founded the EMBL outstation at DESY Hamburg. He was elected to the Royal Society in 1981 and is a member of a number of scientific academies.

John Finch is a retired member of staff of the Medical Research Council Laboratory of Molecular Biology in Cambridge, UK. He began research as a PhD student of Rosalind Franklin's at Birkbeck College, London in 1955 studying the structure of small viruses by x-ray diffraction. He came to Cambridge as part of Aaron Klug's team in 1962 and has continued with the structural study of viruses and other nucleoproteins such as chromatin, using both x-rays and electron microscopy.

Tags: British Biophysical Society, Nature, David Phillips, Robley Williams, David Baltimore, Joe Butler, David Zimmen

Duration: 4 minutes, 46 seconds

Date story recorded: July 2005

Date story went live: 24 January 2008