February 17, 1951.

Dr. Melvin Cohn,
Institut Pasteur,
Paria, France.

Dear Dre Cohn:

Thank you very much for your courtesy in sending the MS. copy of your
paper with Jacques on the ©e cold ML galactosidase. This favor, together
with various hints in Monod's recent, but infrequent, letters, and with
other less reliable sources, lead me te the inference that I was remies
in not having done the same on the occasion of writing my own paper on the
K-12 enzyme. I am sincerely sorry about this embeaion; if I had known that
you had also had a rather parallel centribution in the works, I would cer-
tainly have made a point of it. May I however point to two circumstances:

a) my own paper was written in considerable haste, just as I was packing
for a long trip to California. The MS was mailed as we were leaving town,
and proofs were corrected se we were driving back through Nevada or Utah.

I was never very satisfied with the ns., especially with ths rather sloppy
detaile on lactatic activity, and on the extent of lytic activation, but I
have been asked by several people to domment the INPG technicue, and it
would have taken a great many more months to fron out those details. Another
unfortunate consequence of this haste was the loss of a paragraph which
literally just slipped out during the typing referring to the Nonod-Torrian-
Gribetz note on lactase. I did net see the Am. Inst. Pasteur account until
much later. But more important, I had long since written to Moned about
most of the detaila, especially the im -activation effect. Not having heard
that you had been working along similar lines, I had no reason to suspect
that you would have any special interestg in a subject which fie not at first
sight of much general interest.

Ag you might wmsoect, I read your paper with sone enthueiaem. I shall
look forward especially to seeing the details of your immmochenmical anelysis,
which appears to be approaching a remarkable contribution. Lately, I have
been isolating a number of distinct E. coli strains which csn be crossed
with K-12, and each other. Some of these have distinct sometic antigens, and
I had hoped to fellow up an immunclogical analysis to determine whother they
might not have serologically distinguishable lactases. It might be possible
then to determine whether the determination of enzym:tic and serelogical
specificity might be genetically separable. Your demmnstration of the cross-
reactivity of Serobacter and two colf lactases in somewhat of a damper, however.
At any rate, 44 is quite apparent that we shall have to lean quite heavily
on your exciting werk in this area.

A word about the ms. iteelf. (let: on p.9 you have a reference to tableII
which should read III}. One of the ways in which I ran into the Na activation
came from the fact thet ethanolamine (in -EA-citrate buffer) was markedly in-
hibitory at quite low concentrations. Is it vossible thet the tri~ethanolamine
4s also inhibitory, but that its effect is already maximal (4.2., the competition
with H” ¢ already complete) in the concentrations you used in your buffer
teste. This could shew up by differing sensitivity to K or Na in different
TEA buffer concentrations.
One of the most puzzling features of the ionic activation is the nature
of its effects on the enzyme kinetics (p. 385 of my paper), which suggested
a partialy overlapping of the adsorption site with that of the substrate.
This picture which I proposed as a merely formal one is given considerable
substance by your discovery that the nature of the cubstrate determines
the kind of response. I could net find in your data the mmans to distinguish
between effects on V, and on K,. M/600 ONPG is eceens Berea ny

a
K-12 enzyme in Teabien ionic environments, whereas M/18 lactose ost,
will be under most conditions. 4s part of a general study on thia curious
phenomenon, one of the graduate students at the enzyme institute is studying
the ionic effects on K, (measured as K, against ONPG). I will forward his
findings as they come up. To date, he has been purifying a batch of enzyme
(from 150 g. dry celle - which I understand is quite puny compared to the

output of the bactogen), and claims that cold methanol pptn. is a superior
technique ¥

A first teaction might be that duplication such as ours might be unfor~
tunate, but after a little reflection, I am convinced that this is not so,
especially because of the diversity of material and approaches used. I would
be prepared to discuss any suggestions you might have for planning which
lines of work should be followed in the different laboratpries, but I think
this is probably less important than a frequent and honest exchange of
information. I will admit that this 1s likely to be lopsided, because my
facilities are relatively quite limited, and I can spend only a part of
my time on enzymological problems (for which reason it is, of course, the
genetic aspects which are of closest concem).

Probably, 1 have already mentioned (to Monod) a mutation leading to the
constitutive production of galactosidase. For the present, this seems to be
allelic to the Lac, mutation (which blocks adaptation to lactose but not to
alkgl galactosides’ ). Neolactose (altrose-galactoside) does not provoke
lactase, but is attacked readily by it, again separating the adaptive res-
ponse from the enzymatic attack itself. (imtxnetxutengingxthetxneninctess
mightxeembinaxniihxthaxenz This property of neolactose was the basis of the
original isolation of the constitutive mutation. Gmkiaxgemm Cat cells
grown on glucose are optimally adapted to ONPG but not to galactose, so that
I doubt if it can be a matter of intracellular production of self~adapting
galactosides. We are twa studying now, but unfortunately only slowly, the
interattion of Cat‘ with other genntypes, in hybrids and in mixtures. I would
Bive a good deal te have a system in coli K-12 which adapts a little more
rapidly so that something sould be said of rates of adaptation; from this
point of view, Stenier's enzymes are much more satisfactory (aromatic oxidases
in Pseudomonas). We found this summer that adaptation could be prevented
there with uv, without affecting enzyme activity (his material is not affected
by DNP or azide!). The UV-inhibited calls were sueceotibae to photoreactivation,
showing merely that the photoreactivable process is actually, in some senses,
a terminal one. Ye thought that adaptation was more sensitive than viability,
but the patent clumping of the cells makes this rather uncertain.

With Deutsch on leave (if only that?), there is a certain hiatus in immmo-
chemistry on this campus. How would you assess the chances of viskt back
here on your part?~ or are you planning a more permanent emigration?
Meanwhile, Alain (Bussard) has been showing me and devloping a few tricks
in paper electrophoresis. *xcept that thie technique readily shows the
heterogeneity of K-12 lactase (aseayed on the paper by spraying @IPG),
nothing has come of it yet.

Ces fap (Boil, 9 Bete) Sincerely,
PS: Do yéu want the ms back? Joshua Lederberg