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Finding the origin of the assembly of TMV
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Finding the origin of the assembly of TMV
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Views | Duration | ||
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31. TMV: the biological role of the two-layer disc | 83 | 05:07 | |
32. Producing a phase diagram for the A protein | 67 | 03:06 | |
33. TMV: turning the disc into a helix | 76 | 05:00 | |
34. Finding the origin of the assembly of TMV | 94 | 04:45 | |
35. TMV: the direction of assembly | 72 | 05:01 | |
36. Work on the structure of tRNA | 173 | 05:17 | |
37. Competition to solve the structure of tRNA | 142 | 04:25 | |
38. Creating modern structural molecular biology | 80 | 02:22 | |
39. 'It was the time for chromatin' | 101 | 04:15 | |
40. The citation for the Nobel Prize in Chemistry 1982 | 188 | 01:08 |
At this stage... I suggested this and I did say this in the lab and told Tony Durham, that we ought to use the disc for the assembly. Now, Joe Butler had joined us by this time... Joe Butler, of course, because Reuben was leaving. He was leaving to... he left, I don't know, when did he join you in Germany?
[Q] '69.
'69, so this was '70, so he'd left by them. So he... and I got Joe Butler who was a biochemist, a protein chemist to join us. So I asked... I asked Tony Duhram, who was my research student to do some experiments and make some RNA, probably do it with Joe... Joe Butler and to mix the disc with the pure viral RNA and see if it nucleated assembly, and started the assembly. He wouldn't do it because he'd been told by another research student unit of mine called Peter Gilbert, who was a rather toughy and very determined, who'd been carrying out three-dimensional image reconstruction on discs. And came to the conclusion that the two layers of the disc related by a two-fold axis of symmetry perpendicular to the axis of the disc, which meant that the two layers were back to back. Now... so that meant that... only one layer would be active, but you see in the... if a disc is to turn into a helix then the... the two turns had to be facing the same way, if it doesn't have a two-fold axis for symmetry. Peter Gilbert, being a very persuasive fellow and being a fellow researcher with Tony Durham said that I was, that my ideas wouldn't work at all, that's why he wouldn't do the experiments, he wouldn't do it. So I actually ordered, I don't often have to do this, I ordered Tony Duhram to bring me a solution of acetic acid. And he brought the acetic acid and I poured it, it's one of the few things I ever did with my own hands. I poured the acetic acid into this... into the solution of discs, it all went cloudy instantly. And then we called you and you looked in the microscope and what you could see were large aggregates of dislocated discs on top of each other. And so it was fairly obvious that if protons could do it... turn in changing the structure of the, it was a very facile thing to turn the disc... turn the disc into helix, just like... just like that. And the... And so... but then, Joe Butler was around, so we got Joe to do the experiments, Tony carried on with doing other things as well. This showed clearly that this was the nucleating agent, so then... we... we published a paper on the... on the discs, you and I and Tony Durham about the discs dislocating and forming helices. And over some time they annealed into a perfect helical rod, and so then I got Joe to make some... RNA and asked him to... he'd just joined, to just see if the discs would nucleate the assembly of the protein. So we... we added... the idea was we would nucleate the assembly, so we added the... Joe did these experiments, he made a solution of discs, we knew how to make them in the pH7. He added the RNA and within five or six minutes it began to... it went cloudy. You could see it by eye, I mean, of course it had a light scattering so it was clearly working. So I assumed at the time that the... that the agent would be that the protein contains small aggregates, so to start nucleation and the aggregation or rather the elongation as a biochemist would call it or the growth as a physicist would call it would take place by adding one unit at a time. And this turned out not to be true at all because the... the more discs, when... when we changed the concentration of the discs it went faster and faster and made a more or less a complete virus. And so, the discs were being involved not just in nucleation but also in the growth in dislocating. And so the... And so that is something... so we were able to make more or less complete viral particles in about six minutes compared to the 24 hours. Now, the recipes for making, reconstituting the virus out of protein and RNA, if you looked up Frankel-Conrad's papers, they added some salt to the protein and they waited for several hours before adding the RNA. And I... of course it was obvious what this meant, they were adding the salt to form a disc.
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.
Title: TMV: turning the disc into a helix
Listeners: John Finch Ken Holmes
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.
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.
Tags: Tony Durham, Joe Butler, Peter Gilbert
Duration: 5 minutes, 1 second
Date story recorded: July 2005
Date story went live: 24 January 2008