June 5, 1957
2967
COMMUNICATIONS T O T H E EDITOR THE METHYL GROUP OF METHIONINE AS A SOURCE OF Czs IN ERGOSTEROL'
the residue, representing the ring system. I n the second case this relationship is reversed, as exSir: pected if the main incorporation of radioactivity It was reported recently that formate can serve were in Czs. In experiment 2, the ergosterol, obas a source of Czs in the biosynthesis of ergosterol2 tained from incubation of whole yeast with methioor of eburicoic acid.3 We have incubated cell-free nine-methyl-C14, although recrystallized to conyeast homogenates with labelled sodium bicar- stant specific activity, was subsequently found to bonate, formaldehyde, propionate-( 1 or 2)-C14 be impure. However, the dinitrophenylhydrazone and methionine-meth~1-C'~ and have found that of the a#-dimethyl-butyraldehyde was carefully more C14 is incorporated into the non-saponifiable purified before counting. Further degradation of lipids and the digitonin-precipitable sterols from this aldehyde shows that a predominant portion of methionine than from any of the other compounds. the total radioactivity is located in c28. When the Thus, in one experiment 20.270 of the methionine- correction factor suggested by Ehrensvard, et al.,6 m e t h ~ 1 - C 'radioactivity ~ was found in the non- for the combustion of C14HIsto BaC1403is applied, saponifiable fraction, as compared to 1.2% for the observed activity of is in acceptable agreea ~ e t a t e - 1 - C ' ~1.0% ) for NaHC03, 0.4% for form- ment with the figure calculated from the specific aldehyde and 0.5% for propionate. In compara- activity of the a,P-dimethyl-butyraldehyde. Cartive experiments with the same yeast homogenate bon 28 contains 20-30 times the radioactivity of which we ran after the paper of Danielson and the other carbon atoms in the molecule. Such a Bloch2 came to our attention, methionine-methyl- randomization of radioactivity was also observed C14 gave four times as much radioactivity in the by Dauben, et al., in the incorporation of C14 from digitonin-precipitable sterol fraction as sodium formic acid into the analogous position in eburicoic formate-C14. acid. To determine whether the radioactivity is conThe major incorporation of CI4 into CB of ergoscentrated in CB, samples of ergosterol from incuba- terol in our experiments may be accounted for by tions with acetate-l-C14 and methionine-meth~1-C'~ the well-known oxidation of the methyl group of were ozonized according to Hanahan and SVakil.4 methionine to formate in biological systems.e Comparison of the figures obtained with ergosterol However, transmethylation from sulfur to carbon, made from acetate and from methionine by a yeast which has not been observed up to now, cannot be homogenate (Table I, Expt. 1) shows that in the excluded since we find that aminopterin decreases the incorporation of C14 into ergosterol from forTABLE I mate-C14 but not from methionine-meth~1-C'~. DEGRADATION O F ERGOSTEROL-C14 Other experiments indicate that squalene, but not Source of '2x4, counts/mmole zymosterol, is converted to ergosterol in yeast carbon/min. homogenates. Acetate-1- MetbionineExpt.
1"
ZC
Substance
Ergosterol Steam volatile fraction' Residue from steam distillation Ergosterol acetate a,@-Dimethylbutyraldehyde* Acetone' ( C Z ~ , Z ~ , U ) Cza
Cl'
methyl-Cl4
5,540 6,220
3,843 11,480
7,545
655 946
...
(5) G. Ehrensvard, Q. Reio, E. Saluste and R. Stjernholm, J. Biol. Chem., 189, 93 (1951). (6) J. S. Fruton and S. Simmonds, "General Biochemistry," John Wiley and Sons, Inc., New York, N. Y., 1953,p. 714.
WORCESTER FOUNDATIOX FOR GEORGE J. ALEXANDER ALLEN M. GOLD EXPERIMENTAL BIOLOGY SHREWSBURY, MASSACHUSETTS ERWINSCHWENK RECEIVED MARCH16, 1957
...
1,044 165 .. 110 C*ad ... 3,210 a The two samples of ergosterol in Expt. 1 are not directly comparable, since much more C14 was used in the case of acetate. Isolated as dinitrophenylhydrazone. All samples counted after combustion to BaC08, corrected t o infinite thickness. Combustion of CHI, reportedly gives BaCOs of low specific activity.6
...
DIFFUSION OF
0 ' 8
AND OF PROTIUM IN DZO-HZO MIXTURES'
sir:
We wish to report some interesting and unexpected results obtained in this Laboratory in the course of studies on diffusion in liquid systems.2 The data were obtained for 25 =t0.02' by means of the diaphragm cell technique.2 Deuterium first case the steam-volatile fraction, a, /%dimethyl- analyses were made pycnometrically, and those for ~ butyraldehyde, contains less radioactivity than 0l8 by the gradient tube density m e t h ~ d ,after first converting the samples of D2018-H2018to (1) This investigation was supported by the U. S. Public Health H1Ol8. This last was accomplished by vaporizing Service Grant No. C321, an Institutional Grant from the American Cancer Society, The Schering Corporation, BloomBeld. New Jersey and the Jane Coffin Childs Memorial Fund. (2) H. Danielson and K. Bloch, THIS J O U R N A L , 19, 500 (1957). (3) W.G.Dauben, G. J. Fonken and G. A. Boswell, ibid., '79, 1000 (1967). (4) D. G. Hanahan and S. J. Wakil, ibid., '76, 273 (1953).
(1) These investigations were supported in part by the O 5 c e of Ordnance Research. (2) A. W.Adamson and R. R. Irani, Abstracts 130th Mtg., Amer. Chem. Soc., Atlantic City, N. J., September, 1956. (3) A. Hvidt, G. Johansen, K. Linderstrom-Lang and F. Vaslow, Compl. rend. Irav. Lab. Carlsberg, Ser. Chim., 99, No. 9.
