What Mad Pursuit (18 page)

I soon found that my initial mutant had not one, but several, distinct suppressors, all of which mapped fairly close to the original mutant. I decided that I would have to call them all by a distinctive name. I often worked through the weekend, taking Monday off so that our laboratory kitchen (which did all the washing up and also prepared petri dishes for our use) could catch up. It happened that it was a weekend when I needed a new name, and nobody else was around. Mutants were usually called by a letter, followed by a number. Thus P31 meant the thirty-first mutant in the P series, probably produced by proflavin. Unfortunately I could not remember for certain which letters had already been used, so I decided to rename my mutant FCO, since I was quite sure that no one had used my initials to name mutants. The new suppressives were then named FC1, FC2, and so on. This use of my own initials suggested to some people that I must be conceited, but the real explanation was that I have a rather fallible memory.

The new suppressors all seemed like good, nonleaky mutants. So why not, I argued, see if they too had suppressors? And indeed they had. I even went a step further and found suppressors of suppressors of suppressors.

So what was going on? Fortunately we had the right ideas already at hand. Assume that the genetic message was read (to produce a protein) in steps of three bases at a time, starting from one particular point in the message. To make it clearer, let us take an extremely simple message that merely repeats the triplet TAG over and over again:

. . . TAG, TAG, TAG, TAG, TAG, TAG, . . .

the dots indicating that there is message both before and after such a sequence. Commas have been added to show in which “phase” the sequence has to be read. I assumed that this phase was determined by a special “start” signal, somewhere to the left of the stretch shown.

Assume that our original mutant (now called FCO) had added a base to the base sequence. Then, from that point on, the reading would be out of step (out of phase) and thus would produce a nonsense protein, a protein whose amino acid sequence, following the mutant, was completely incorrect, so that the gene product could not function.

Our simple sequence might have become

(The added base has been shown, for clarity, as a C, but it could have been any of the four bases.)

Then, on this interpretation, a suppressor, such as FC1, was the
deletion
of one base at a point nearby. In between FCO and FC1 the message would still be incorrect, being read in the wrong phase, but elsewhere the reading would be normal.

Our example might thus become:

If the altered bit of the amino acid sequence was not crucial (and in this case there was other evidence to suggest this), then the protein would still function fairly well and the double mutant (FCO + FC1) would behave more like a wild type than like a nonleaky mutant.

I therefore labeled all the first set of suppressors—. The next set, the suppressors of the first set of suppressors, we labeled +, and
their
suppressors we labeled—.

I had started these experiments early in May and by now summer was advancing. I had previously arranged to take my family on a summer holiday, almost the first proper holiday we had ever had, since by now my financial position was a little easier. We had rented, for a very small sum, a large villa on the old mountain in Tangier, a town in North Africa, just opposite Gibraltar. Here we lived in splendor, with one Arab servant living in and another coming each day. Odile and our German au pair girl, Eleanora, learned how to shop for food in the Arab market, bargaining, walking away, and so on. Our two daughters improved their swimming on the beach while I usually spent the day on the terrace, in the dappled shade of the palm trees.

On the way to Tangier I attended a scientific meeting. Even in those days scientists were reluctant to go to a meeting unless it was in some interesting place. This meeting was at Col de Voz halfway up Mont Blanc. I reported my preliminary results, which eventually were published as a very brief communication related to the meeting.

After a month in Tangier I went off to the 1961 Biochemical Congress at Moscow, leaving my family to stay at the villa for another week or so. Moscow then was very different from my first visit in 1945, during the war. Now it was summer, rather than the depth of winter, and everything was brighter and more prosperous than in the drab days of wartime. I stayed in a student’s room in the university, where the meeting was held, and got to know some of our Russian hosts. A dominant figure was Igor Tamm, the Russian physicist. The influence of Lysenko, the man who had, for a period, killed genetics in the USSR, was very much on the wane. I sensed that his eclipse was largely the work of physicists like Tamm who had considerable political influence and who could recognize scientific nonsense when they saw it. A number of us were invited to give talks to the biological section of the Russian Atomic Energy Research establishment, something that could not have happened a few years before. We gave our talks in English, but they were brilliantly translated (in chunks, as we went along) by Bressler, a Russian scientist we had already met when he had visited Cambridge. Bressler not only understood what we were saying but in some cases, as I could tell by listening to him, filled out the “references” the speakers were giving, a truly remarkable performance.

The Moscow meeting was made especially interesting because of the results reported by Marshall Nirenberg, then almost unknown. I had heard rumors of these experiments but no details. Matt Meselson, whom I ran into in a corridor, alerted me to Marshall’s talk in a remote seminar room. I was so impressed that I asked Marshall to take part in a much larger meeting, of which I was the chairman. What he had discovered was that he could add an artificial message to a test-tube system that synthesized proteins and get it to direct some synthesis. In detail, he had added poly U—the RNA message consisting entirely of a sequence of uracils—to the system and it had synthesized polyphenylalanine. This suggested that UUU (assuming a triplet code) was a codon for phenylalanine (one of the “magic twenty” amino acids), as indeed it is. I later claimed that the audience was “startled” (I think I originally wrote “electrified”) to receive this news. Seymour Benzer countered this with a photograph showing everyone looking extremely bored! Nevertheless it was an epoch-making discovery, after which there was no looking back.

There was also a measure of social life during the week in Moscow. I enjoyed visiting an old-style apartment, with heavy furniture and a bed behind a large bookcase. Also a more modern one, with a much lighter tone. The owner collected modern Russian art. I was amused to notice Alex Rich demonstrating a strange new American dance to our host, a dance I later recognized as the twist. As Alex’s waist is not very pronounced, the twist, as demonstrated by him, was somewhat less than free-flowing.

I returned to Cambridge. The next step was to do further experiments to validate the ideas that there was some sense in labeling each of our new r
_II
mutants as either + or −. The theory predicted that any combination of the type (+ +)or(− −) would be a mutant. My colleagues and I constructed quite a number of such pairs and they were all nonleaky mutants, as predicted. The simple theory also predicted that
any
combination of the type ( + −) would be wild type, or approximately so. Of course we knew this to be true in some cases, since that was how we had picked up the suppressor in the first place, but many other combinations (of a + with −) had never been tested. These we called “Uncles and Aunts,” since creating them often involved putting together a mutant of one generation with a mutant from a previous generation, but one other than the one it was descended from. I had asked Sydney to see that some of these were tried while I was away but he had other ideas, so I had to do it myself when I returned.

At this point a small complication arose. Some of the (+ −) combinations, predicted to be wild type, turned out to be mutant. We explained these away by assuming that in some cases the small local phase shift between the + and the − produced a “nonsense” mutant. We know now that these nonsense sites were due to a triplet that terminated the polypeptide chain, thus producing a nonfunctional protein fragment. I also realized that this depended on the precise phase of the reading. For a nonoverlapping triplet code there is one correct phase but
two
incorrect ones, so that a combination (+ −), that is, + followed by −, will be locally different from a (− +) combination.

To return to our simple example, a (+ −) combination might be:

and a (− +) combination

The first has GTA between the two alterations; the second has AGT. We showed that our (+ + −) or (+ − + ) failures obeyed this rule, which made us fairly confident our ideas were along the right lines.

Previous to this Sydney had an idea. He reasoned that a (+ +) mutant might backmutate to a wild type. He tried one, but the back mutation must have been too close to an existing one, since he could not separate it. Another, slightly more laborious approach was to construct a triple mutant, of the form (+ + +) or (− − −). According to our ideas, these should be wild type, since the three successive changes in phase should have restored the correct phase, always assuming, of course, that it was a triplet code.

For our simple sequence, an example might be