January 6, 1939
Dear Linus,
I would like to know your views on certain questions concerning protein denaturation.
(1) When a protein spreads to form a film it unfolds. At the same time it becomes
insoluble and its SH groups become available. We would say that it is denatured.
And yet it has certain important properties of the native protein. Gorter has studied
the enzymatic activity of trypsin after spreading. The interpretation of his observations
is, however, ambiguous. Langmuir's exp[eriments] on urease seem pretty clear. While
spread out as a film, this protein acts as an enzyme. In this sense it is not denatured. We regard unfolding as taking place in denaturation and yet here is a
case of unfolding without loss of a certain configuration. I feel the explanation
of the apparent contradiction is as follows: when a protein spreads unfolding takes
place in a very orderly manner. Langmuir came over here on day and demonstrated some
of his exp[eriments]. As a protein spreads it forms certain visible patterns. The
same protein always forms the same pattern; each protein forms a different and characteristic
pattern. The whole film forms a continuum, bonds holding everything in one piece.
When a protein unfolds at a surface it seems possible for it to do so without passing
into a multitude of random configurations. The configuration of the protein in the
film may in certain respects be very similar to that of the original native protein.
It may therefore still act as an enzyme. At the same time it is insoluble and has
its SH groups available. It has some of the properties of native protein and some
properties of denatured. The controversy about whether or not it is denatured is
in a certain sense surely verbal. What do you think of this?
(2) Another point about configuration and specific properties. When the layers of
a protein fall apart from each other to assume random relations to each other, certain
specific properties of proteins are lost - as mentioned in our paper. Other properties
distinguishing one protein from another will remain. The amino acid contents of proteins
vary and the sequence of amino acids in the chains vary from one protein to another.
Even after unfolding these differences will persist and manifest themselves in certain
properties. There is good evidence that antibodies are proteins. These proteins lose
the power to act as antibodies on denaturation in the same way that Hb and the enzymes
lose their properties on denaturation. Unfolding (with subsequent random arrangements)
is enough to denature antibodies, Hb and enzymes. An antigen is a foreign protein which when injected into an animal gives rise to antibodies.
There is some evidence (by no means conclusive) that when an antigenic protein is
denatured in the same way as Hb, enzymes etc. are it loses some of its specific properties,
but not all of them. It certainly is possible on our views that certain properties of a specific nature would remain after denaturation
and there is experimental evidence that this is so. In a certain sense, denaturation
will be complete only when the protein is finally resolved into its constituent amino
acids. There do, however, appear to be certain definite steps in the loss of specific
configurations and our paper was concerned with one of the first and apparently most
definite steps.
(3) Have you seen the paper by LaMer in Science of Dec. 31st? Please let me know what
you think of this. It seems to me that LaMer does not consider the view that denaturation
takes place only after many bonds are broken - after a considerable number of steps
(and one after the other) have been taken, so to speak. What do you think of Eyring's
views? What about the meaning of S, the steric factor, in the velocity equation? What
about the role of collisions in denaturation?
(4) I have made some progress in understanding the coagulation of myosin in muscle.
You may remember I told you about comparing the effect of heat on the contraction
of muscle (in which myosin coagulates) and on the coagulation of isolated myosin.
Frog muscle is caused to contract by raising the temperature to 37 degrees and isolated
frog myosin coagulates at 37 degrees. But the rate of this latter process is very
slow. I now find that if myosin is treated by NH4Cl or LiCl (and the excess or free salt carefully washed away) that the myosin has
become sensitized. It is now exceedingly unstable. At 37 degrees it is coagulated
with very great speed. Heretofore I have prepared myosin by using KCl. After treating
myosin with LiCl or NH4Cl the ppt. of myosin can be washed for at least 10 hours with constantly renewed
dilute KCl and still this myosin appears sensitized when compared with myosin that
has never been treated with anything but KCl. How much LiCl or NH4Cl remains attached to the myosin I do not yet know. The instability of the myosin
shows itself in other ways too. If ordinary (KCl) myosin is dried it also loses its
solubility - it cannot be redissolved. If a surfusion of this myosin is frozen at
-10 C it slowly in the course of weeks loses its solubility. It is presumably being
dried by freezing away its water. Myosin that has been treated with NH4Cl loses its solubility after it has been frozen for several hours. The possible bearings
of all this on muscle - In the muscle fibre K is the common cation and it is located
in the same place in the fibre as is myosin. During contraction K leaves the fibre,
passing into the blood, and returning during rest. At the same time (approx) myosin
becomes insoluble. Also at the same time the fibre becomes opaque. If gels (the myosin
in both are still soluble) of myosin of the same concentration are compared (one of
the gels having previously been treated by NH4Cl) it is striking how much more opaque the NH4Cl treated myosin is than ordinary KCl myosin. It looks as if K stabilizes myosin
in the muscle fibre and that in activity NH4 or something like it comes in contact with myosin and sensitizes it. All this is
quite new and incomplete, but the observations I have just described to you are, I
believe, probably sound. Perhaps this has been too incompletely described to be comprehensible.
If you still are in the East, it would be the greatest pleasure for me to see you
again. I would be quite willing to travel a bit for a talk with you.
In the meantime, yours as ever,
Alfred Mirsky
O, by the way, I was talking with Gasser about you the other day. He referred to you
as a "wizard." I hope you don't find being one a burden.
