VIII Nov. 16, 1937
Dear Dr. Pauling:
This idea of writing a weekly letter may make the letters . discuss work weakly [sic]. Taylor says that I spend so much
time writing you that I do not have time to get any work done. You have not fired me yet, but that may be because that you
are too busy.
The apparatus that I got with Webb Tuesday at Corona is in pretty good shape for developing the dialyzing apparatus, requiring
only a motor with a good reduction gear and the glass parts, but Webb has not yet set it up completely. I was going to
give him the motor from my rotor-stirrer but I have used it a good deal this week, and may want to go on with it. Webb
has gone today for more blood although he has a stock of two liters from our last visits that is not used. Ithink [sic]
that I told you a little last week about our plans to get the isoelectric point (rather important to determine) and simultaneously
get some data on the effect of salts on the pH and solubility of globulins. I think that we have worked out a pretty good
method that will not be too difficult to put into practice. We will take a suspension of dialyzed (isoelectric [handwritten
annotation: "net charge zero"] and isoionic [handwritten annotation: "equal amounts H+·OH- bound"]) hemocyanin, and titrate with various salts like NaCl and NaNCS and determine simultaneously pH and amount of protein
that has gone into solution. With hemoglobin, NaCl affects the pH differently on opposite sides of the isoelectric point,
decreasing it on the alkaline side, increasing it on the acid side (Cohn, paper in the JACS Mar. 1937). This is probably
a normal [handwritten annotation: "salt"] effect, and although large, is in the direction of expected effects of ionic strength,
or of appreciable absorption of both Na+ and Cl-, not greatly different [handwritten annotation: "in amounts"]. With KNCS the pH increases on addition to HB+ solution on both sides of the isoelectric points, pointing to absorption of NCS- far greater than of K+, the effect partially-offset by taking on of protons from the solution. Of course with hemocyanin there is the added
effect (sofar as I know as yet unexplained) of salt in bringing the protein [edit: "globulin"] into solution, but a treatment
of the type you gave to Carpenter's experiments on gelatin may be significant. The order of ions in the lyotropic series
in bringing globulins (or any colloid) into solution is the same order that prevails in salting out, which indicates strongly
that the effects are due mainly to the same forces.
Taylor has been working very hard the last few weeks, and has three complete determinations of the absolute moment of hemoglobin
(whole blood) worked out, involving the ordinary (but very precise) magnetic measurements as well as both oxygen and carbon
monoxide capacities. I got blood for him twice last week and will have to go twice this week. I determined some time-
ago that there is absolutely no change of Δw of Hb on laking corpuscles with saponin, but you may recall in our work that
the moments of ether-laked Hb solutions C and D may have been lower than moments of whole bloods A and B. Taylor and I are
laking with toluene, and find it satisfactory on 24-48 hours standing in the refrigerator. The method used by Mirsky based
on work of Henderson (1921) (vol. missing from biol. libr.) involved diluting the washed corpuscles with an equal volume of
water (which in itself brings about considerable laking) and then allowing to stand with toluene overnight. Taylor could
find no intact cells and very few 'ghosts' microscopically in [edit: "a solution from"] some corpuscles [handwritten annotation:
"without water"] that I had shaken with toluene, allowed to stand 48 hours, and then centrifuged. This improvement allows
us to obtain the more concentrated solutions, and is free from objections inherent in peroxides or peroxide-formation in the
presence of ether. Of course we shall check the ether-laking but for future work probably use the toluene.
Taylor is interested in getting the best value of the absolute moment of Hb, and finds considerable difference (up to 5%)
between concentrations calculated on the basis of carbon monoxide capacity and oxygen capacity. (This would be predicted
from the work of Barkan.) The moment calculated from magnetic weighings and the former capacity is accidentally very close
to the value that we used (within several hundredths of a Bohr magneton) so we will not have to make a serious correction
in published moments, since they should be referred to this standard rather than to one based on oxygen. We shall have
to think hard, when all of the data are ready, to explain the probable source of some of the apparent differences.
Taylor is, by the way, waiting to hear from you about what to start next, as he is near the end of this work.
I conceived the idea of fortifying Dodson's work on the magnetic titrations of HbO2 with Na2S2O4 with similar work based on the following reaction:
2 Hb + 2 NO-2 + S2O=4 = 2 HbNO + 2 SO=3, adding standard nitrite to Hb in the presence of excess S2O=4. In two carefully made titrations I ran out of reducing agent before I reached the end-point, but the curves had a very
nice linear early portion. After adding an excess of nitrite I could get the final Δw by adding another portion of hyposulfite
(I prefer this name now to hydrosulfite). The only difficulty is that, with identical initial volumes of Hb+ reduced, etc., the slopes if the linear portions differed appreciably. The slope for the second titration was less, which
may indicate that the nitrite-had decomposed somewhat, although that does not seem too likely to me. The chief objections
have to our earlier work with Dodson are that the hyposulfite decomposed during the course of the reaction, which would lead
one to expect a curvature of the opposite nature to the one expected, and that transformation of oxyhemoglobin to ferrihemoglobin
occured spontaneously to a large enough extent to cast doubts on the validity of the work. Palmer's uncle-in-law, Baumberger
at Stanford, told me once of the great success he had once reducing oxyhemoglobin at a constant rate with bacteria (yeast,
I believe works too). If I could get a culture that would eat fast enough that the reaction was over before multiplication
of the beasts became serious, this would be a good possibility. I believe that in the presence of organisms ferrihemoglobin
would be reduced as fast as it formed. I shall write him for advice. We sent him a slug of sodium azide recently for his work,
and I have met him several times.
I have a paper half written (and typed that far by GraceMary) which I want to submit to you for approval on the acid-base
functions of the heme of hemoglobin and myoglobin. As long as I had all this material worked up for seminars, I thought that
it would be advisable
to see how it would look written up, but I do not intend to waste a
great deal of time on the matter because there is no experimental work of mine in it. It consists merely of a thorough correlation
of all of the work I can lay hands on, interpreted from a structural viewpoint. There has been a lot of uncorrelated work
done in this field, and a fair amount of wrong mathematical treatment, but of course, a fair amount of correct handling of
isolated parts of the problem have appeared. I shall leave the matter up to you.
I gave up my projected trip to Davis and Deep Springs because my passengers backed out. I found that they were having only
the morning session at Davis and thought that the place was too far away for a trip for so short a meeting.
Mrs. Pauling said that you are staying at the Telluride House. I hope that you don't get tellurium poisoning there. From
what she says both lecture and book are progressing wonderfully, for which I am very glad. However, do not forget to take
a little time off from work for relaxation once in a while.
[handwritten annotation: "Yours, Charles Coryell"]