The mechanism which Monod proposed is that.. well, Monod really is a Platonist, it's a symmetry mechanism. It said, if one sub-unit changes its confirmation to the other, then all the other sub-units have to come along to make it symmetric. So then you see already that a Monod mechanism consists of at least three steps. One step is conformation one... which is, let's say has a high affinity to its ligand... so there are four sub-units, so four ligands of four substrates might bind to the enzyme, so you have a series of binding steps, coupled binding steps. Then you have the other confirmation of different, lower affinity, again four steps of binding, but the enzyme can be either in that state with all its sub-units or in the other states. So you have a co-operative transformation between the high affinity and low affinity state. So you must find at least three types of rate constant, binding in the high affinity, binding in the low affinity, confirmation change among them, and that's exactly what we found. Kaspar Kirschner did this study. There is one reaction in the range of microseconds, then there's another reaction in the range of 10 to 100 microseconds, and there's a third one in the millisecond range and one showed the concentration dependence of binding high affinity, the other low affinity, and the confirmation changes of first order reaction. So this was a big success and Monod came here and he was happy.

[Q] So I understand that to measure this reaction was only possible in your laboratory with a relaxation method.

That's right.

[Q] I remember, I think it was a T-jump.

It was done with a T-jump. And this T-jump method works down to a microsecond and that was indeed the shortest time in that reaction.

Well, and then there was Koshland. Koshland proposed the different mechanism, which I might explain also. He had the idea of induced fit, which means... there is no symmetry among them.  If the first ligand binds, it induces a fit, and then increases the affinity of the next sub-unit, and if the second binds it still increases it for the next so that you have the successive binding with increasing affinity. Well it turned out that this first mechanism did not comply with the Koshland idea, but later on we found many other systems which followed the Koshland rather than the Monod mechanism.  And we came up at that time with the very general mechanism in which you can combine both, and at that time also the biochemists really realised that this method really could give them new insights also into the reaction of the living part.