Download Metabolism of Resistant Mutants of

Metabolism of Resistant Mutants of Streptococcusfaecalis
III. The Action of 6-Mercaptopurine*t
M. EARLBALIS,VALIAHYLIN, M. KATHARINE
COULTAS,ANDDORRISJ. HUTCHISON
(Laboratories of the Sloan-Kettering Institute for Cancer Research, New York, N.Y.)
The compound 6-mercaptopurine (6-MP) has
been shown to possess definite antitumor (13)
properties and to inhibit the growth of several
microorganisms (14-16). This inhibition has been
reversed by several purines (14-16), and it is
possible that the antitumor action is also con
cerned with purine metabolism. The present exper
iments were undertaken in an attempt to elucidate
the action of 6-MP by studying its effects on
bacteria which are resistant to it. Several such
strains have been isolated and characterized (17),
and certain aspects of their metabolism have been
described (4, 17). In the present study the followbig mutants have been used:
SF/O
Parent (ATCC No. 8043)
SF/MP
Resistant to 6-MP
SF/MPcc Resistant to 6-MP, isolated inde
pendently of SF/MP
SF/MP/A Double mutant, resistant to 6-MP
and A-methopterin
SF/DAP
Resistant to 2,6-diammopurine
SF/A
Resistant to A-methopterin
SF/A/O
Partial revert of SF/A
SF/A/MP Double mutant, resistant to Amethopterin and 6-MP
MATERIALS
AND METHODS
Reagents.—The6-MP-8-C" was obtained from the Southern
Research Institute, Birmingham, Alabama, and the sodium
fonnate-C14 from Isotopes Specialties Co., Burbank, Califor
nia.
Bacteria and media.—Theisolation, maintenance, and char
acterization of the strains of Streptococcusfaecalia have been
described elsewhere (16, 17). The standard inocula used in
these experiments were grown and prepared according to the
following regimen: Two successive transfers were made into 5
ml. of liquid medium identical with that on which the culture is
carried; the third transfer with the use of saline-washed inocu
lum was then made into 5 ml. of an unlabeled equivalent of the
medium which was to be used in the labeling experiment. The
* A preliminary report of this work has been presented (3).
t This investigation was supported in part by funds from
the National Cancer Institute, National Institutes of Health,
Public Health Service (Grant # CY3190, C2699), and from the
U.S. Atomic Energy Commission—Contract # AT(301)-910.
Received for publication November 27, 1957.
inoculum used was one which gave approximately 8 X 10'
cells/ml. In each experiment, 400 ml. of a folie acid-(l m¿tg/ml)
or thymine-(l jug/ml) supplemented purine and pyrimidinefree medium (F-PP) was further supplemented with the labeled
purine at a concentration of 3.8 X 10~~*
M. This medium was
sterilized at 121°
C. for 15 minutes; if the labeled compound
was heat-labile, it was omitted from the medium, sterilized by
filtration through an ultrafine sintered glass filter, and added
aseptically to the cooled medium. Just prior to inoculation, a
K'-ml. aliquot was removed aseptically from each flask to serve
as a titration blank. The flasks were then inoculated so that there
were approximately 8 X IO6cells/ml and incubated at 35°C.
for 18 hours. At the termination of the incubation period, 19
ml. were removed from each flask, and the rest of the cells were
harvested by centrifugation. A 5-ml. aliquot was handled
aseptically, since it was to be used as the inoculum for a series of
controls to determine the resistance and purine requirement for
the mutants after the growth under the above conditions. A
2-ml. sample was used to determine the amount of growth by
turbidity measurement on a Coleman Junior Spectrophotometer, and a 12-ml. sample was used for titer determination.
Controls on A-methopterin and 6-MP resistance as well as
purine requirements were made on each culture for each experi
ment; the saline-washed suspension prepared from the 5-ml.
aliquot removed from the experimental flasks was used as such.
The basal medium for the resistance controls was the one sup
plemented with PGA at 1 mjug/ml, while the response to adenine, guanine, hypoxanthine, and xanthine was determined in
this medium and a medium supplemented with thymine at 1
Mg/ml rather than the PGA.
The controls were set up in 13 X 100-mm. tubes which con
tained 2 ml. of medium. These tubes were sterilized at 121°
C.
for 8 minutes, cooled, and inoculated with a saline-washed sus
pension diluted so that each inoculated tube contained 8 X 10s
cells/ml. Turbidity measurements were made after incubation
for 18 hours at 35°C.
Isolation of nucleic add purines.—Theharvested celb were
washed successively with cold trichloroacetic acid, alcohol, and
ether. The washed cells were treated at 37°C. for 24 hours with
1 ml. of l N sodium hydroxide per 180 mg. of bacteria (20).
The degraded pentosenucleic acid (PNA) was separated from
the deoxypentosenucleic acid (DNA) by acidification with
HC1 and trichloroacetic acid, followed by the addition of 1.5
volumes of etbanol. The DNA was collected by centrifugation.
The individual purines were isolated (8) by hydrolysis of the
nucleic acids, precipitation of silver purines, regeneration of the
purines as hydrochlorides, and separation of the individual
purines by paper chromatography. The radioactivities were
determined as has been previously described (7). Infinitely thin
Sims on aluminum planchéiswere measured in a Geiger-Müller
flow counter with helium-isobutane gas. All the plancheta con
tained sufficient radioactivity to result in measurements of at
least twice those of the background, and the activities were
determined to within standard errors of less than 5 per cent
(18), except where noted.
440
Downloaded from cancerres.aacrjournals.org on June 11, 2017. © 1958 American Association for Cancer Research.
BALISet al.—Streptococcus faecalis and 6-Mercaptopurine
RESULTS
Table 1 shows the effect of 6-MP on the ability
of the wild strain and seven mutants to synthesize
nucleic acid adenine and guanine from exogenous
xanthine and hypoxanthine. Xanthine is the only
purine which all the strains are known to utilize
for this purpose (4,17). 6-MP had some inhibitory
effect on the conversion of xanthine into nucleic
acid guanine, but greatly depressed its incorpora
tion into nucleic acid adenine in all the strains
except the two single mutants which are resistant
to its growth inhibitory effect, SF/MP and SF/
MPcc. If, however, hypoxanthine was used as the
labeled precursor instead of xanthine, 6-MP caused
a clearly marked depression of the incorporation
of the exogenous purine into both the nucleic acid
adenine and guanine. This depression was very
much greater in the two 6-MP-resistant single
mutants than in any of the other strains. In both
these experiments, effects produced by 6-MP on
the incorporation of the exogenous hypoxanthine
or xanthine into nucleic acid purines were of the
same order of magnitude in both the DNA and
UNA.
The effect of 6-MP on the possible utilization
441
of inosinic acid by the organisms is shown in Table
2. Labeled inosinic acid was not available, so the
method of reciprocal labeling was employed. The
fraction of the nucleic acid purines which was
synthesized de novo is the ratio of the relative
specific activity (RSA) of the nucleic acid purines
obtained from bacteria grown in the presence of
the potential precursor to the RSA obtained from
the control bacteria where both the media con
tained sodium formate-C14 (7). In Experiment 3,
Table 2, are shown the control values for a series
of determinations. In Experiment 4, inosinic acid
was present, and, since there was no reduction
in the amount of formate used for the synthesis
of the nucleic acid purines, it is clear that exoge
nous inosinic acid did not serve as a purine source
for these bacteria. In Experiment 5, both inosinic
acid and 6-MP were added, and certain of the
mutants showed a very large drop in the amount
of formate incorporated into the nucleic acid pu
rines. This might be due to the utilization of either
6-MP or inosinic acid as a purine precursor. ¡How
ever, the addition of 6-MP caused a marked reduc
tion in the incorporation of formate-C14 by five
of the seven strains examined. This indicated that
TABLE1
EFFECTOFO-MERCAPTOPUHINE
ONTHEUTILIZATION
OFEXOGENOUS
PURINES
FOBNUCLEIC
ACIDSYNTHESIS
HIP.t
EXP. 1
AdeninePNA
AdeninePNA.77.19.59.11.64.83.52.54DNA.74.21.81.14.951.20.67.74GuaninePNA.68.16.40.09.68.89.60.57
STRAIN
DNA.03*
.03*1.40
SF/O
1.10.03*
SF/MP
.03*.77
SF/MP/A
SF/MPcc
.58.10
SF/DAP
.11.05*
.07*.03*
SF/A
.03*.01*
SF/A/0
.02*GuaninePNA.81.82.561.001.001.22.58.60DNA1.30.75.881.30.97.63
SF/A/MP
The bacteria were grown in media which contained 1 X 10~3moles/liter of 6-mercaptopurine and 3.8 X 10~*
moles/liter of the labeled purine. In Exp. 1 the labeled precursor was xanthine; in Exp. 2 it was hypoxanthine.
The activities of the isolated purines have been divided by the activities of the corresponding purines from con
trol bacteria grown identically, except for the omission of 6-mercaptopurine.
* Error of radioactivity assay < 15 per cent (18).
TABLE 2
UTILIZATIONOFLABELEDFORMATEBY S. faecalis
EXP. 8
EXP. 4
EXP.«le
«AdeninePNA8*104*0.1*DNA8*104*0.1*GuaninePNA8*104*0.1*
Adenine
Guanine
Adenine
GuanineAdenine>NA1918ID•1tttt18PNA6188*18103*0.1*DNA9178*U148*0.1*GuaninePNA7178*U103*0.1*DNA7178*18
PNA DNA PNA DNA
PNA DNA PNA DNA
STRAIN
21
22
Õ2
18
«2
23
28
SF/O
20
18
19
18
18
18
SF/MP
17
SF/MP/A
17
17
17
17
17
U
•i1«
SI
18
SF/MPcc
to
tt
88
22
SF/DAP
tt
tt
tt
tt
tt
tt
2Õ
tt
SF/A/O
tt
tt
M
10 U
18
1*
SF/A/MP
1»
1»
Values reported are relative specific activities.
(RSA) =- c0110^/"""/1110^ °*isolated baae
IQQ
counta/ min/ mole of precursor
Exp. 3, medium contained 20 ¿ig/ml«odiumformate-C'14.
Èxp.. 4,, medium contained 20 /ig/ ml sodium formate-C14 plus 3.8
X 10~*moles/liter of inosmic acid..
Ezp. 5, medium contained 20 //ir/ml sodium formate-C14 plus 3.8. X 10~*moles/liter
~
of inoainic acid and 1 X 10~*molea/Iiter of 6-mercaptopurine.
Exp. 6, medium contained 20 tai/nú sodium formate-C14 plus 1 X 10~* moles/liter of 6-mercaptopurine.
* Error in radioactivity
assay < IS per cent (18).
Downloaded from cancerres.aacrjournals.org on June 11, 2017. © 1958 American Association for Cancer Research.
442
Cancer Research
these strains, instead of synthesizing purines de
novo, were utilizing either the 6-MP or the inosinic
acid. Experiment 6 showed that, with at least four
of the strains, 6-MP produced a similar depression
of formate incorporation in the absence of inosinic
acid. In these strains, therefore, the 6-MP and
not the inosinic acid was being utilized as a source
of purines for nucleic acid synthesis. It is thus
probable that in the experiments in Table 2 none
of the strains used inosinic acid.
Two strains in which incorporation of the formate-C14 into nucleic acid purines was not de
pressed in the presence of 6-MP were the two
resistant single mutants—SF/MP and SF/MPcc
(Table 2, Experiments 3 and 5). Confirmation of
Vol. 18, May, 1958
it did so to a lesser extent than did any of the
other strains except SF/MP. At the other extreme,
SF/A/MP utilized 6-MP almost exclusively as a
source of nucleic acid purines unless provided with
an alternative purine in the form of xanthine or
hypoxanthine. Comparison of the results in Table
3 with those in Table 2 shows considerable con
firmation. For example, in the case of SF/A/MP,
Exps. 3 and 4 show that, in the presence of
6-MP, incorporation of formate into nucleic acid,
adenine, and guanine was reduced to 0.1/19 or
0.5 per cent of its control level, suggesting that
about 99.5 per cent of nucleic acid adenine and
guanine were derived from the 6-MP.1 Table 3,
Exp. 8, shows by direct measurement that about
TABLE 3
INCORPORATION
OFEXOGENOUS
O-MERCAPTOPURINE
INTONUCLEICACID
PURINESIN MEDIUMCONTAINING
FOLICACID
mjjg/ml)EXP.
9AdeninePNA625*5024560474DNA685*6024880281GuaninePNA416*2920434048Ex».AdeninePNA453*S3172435344410GuaninePNA02»0IS10779Values
8Guanine
7Adenine(1 EXP.
AdenineSTBAIN
PNA54
DNA017971
PNA
PNASF/0SF/MPSF/MP/ASF/MPccSF/DAPSF/ASF/A/OSF/A/MP6190£0976486DNA
PNA
DNA
7085GuanineDNA 7083
86Exp.
as:Relativespecificactivity(RSA)counts/min/molemiint.s/min/inoie
reported
purinexoffenous
Exp.
Exp.
Exp.
Exp.
7, contained
8, contained
9, contained
10, contained
100 X
100 X
100 X
3.8 X
> U I I II L3/ 111I1J/
1LHJ1C
CAUgCUL/1»
eisolated
Dunne100.
JJU11LIC
lO-'MO-MP-S-C" and 3.8 X 10"* Mhypoxanthine.
lO^MO-MP-S-C".
lO^MO-MP-S-C" and 3.8 X 10~6 Mxanthine.
10-*M6-MP-8-C» and S.8 X IQ-" Mxanthine.
* Error of radioactivity assay < IS per cent (18).
the ability of the other strains to use 6-MP as
a source of nucleic acid purines is afforded by
experiments in which 6-MP itself was the labeled
precursor, as may be seen in Table 3, experiments
7 and 8. 6-MP was extensively utilized by strains
SF/A/O and SF/A/MP as a precursor for nucleic
acid purines, and the extent of this utilization was
not greatly modified by the presence of hypo
xanthine. Furthermore, even in the presence of
xanthine seven of the eight strains studied incor
porated labeled 6-MP extensively into their nucleic
acid purines, and this incorporation was quite
substantial even if the unlabeled xanthine and
labeled 6-MP were present in the medium in equal
concentrations (Exps. 9 and 10 in Table 3). The
only strain which did not incorporate 6-MP ex
tensively was SF/MP, one of the single mutants
which is resistant to 6-MP. In this connection it
will be noted that, although the other 6-MPresistant single mutant SF/MPcc utilized 6-MP,
79-86 per cent of the nucleic acid purines were
derived from this source.
It is interesting to note that, throughout these
experiments, irrespective of the precursor used, the
RNA and DNA had, within the limits of experi
mental error, the same specific activities (5). Fur
thermore, the effects of 6-MP were the same on
both nucleic acids.
DISCUSSION
In a previous paper the effects of 6-MP on the
synthesis of purines de novo and the conversion
of one purine derivative into another in Lactobacitttis casei were described (6). It was suggested
that, in this organism, the enzyme which normally
converts hypoxanthine and other purines to their
nucleotides also converts 6-MP to 6-MP nucleo1The basis for this calculation has been discussed in more
detail elsewhere (7).
Downloaded from cancerres.aacrjournals.org on June 11, 2017. © 1958 American Association for Cancer Research.
BALIS et al.—Streptococcus
tide2 and that the latter competes with inosinic
acid and prevents its conversion to adenine and
guanine derivatives. If it is assumed that 6-MP
acts on S. faecalis by the same mechanism, the
simplest type of mutant to be expected is one
which cannot convert 6-MP to its nucleotide.3 It
would further be expected that such a mutant
would also be unable to convert other purines to
their nucleotides. SF/MP seems to be a mutant
of this type. It has already been shown that it
requires xanthine for optimal growth (17). It can
convert hypoxanthine, adenine, and guanine to
their nucleotides, but to a slight extent. This latter
process is inhibited by 6-MP, but it makes such
a small contribution to the economy of the cell
that its inhibition has no effect on growth. The
cells can still utilize exogenous xanthine exten
sively (4, 17), and the amount of 6-MP nucleotide
formed is presumably too small to inhibit utiliza
tion of the large amount of xanthine nucleotide
formed.
The two single mutants which are resistant to
6-MP, SF/MP and SF/MPcc, do have certain
common characteristics which distinguish them
from the other strains: 6-MP depresses their ca
pacity to utilize hypoxanthine, but not xanthine
or formate, as precursors of nucleic acid purines
(Tables 1 and 2). They differ, however, in that
SF/MPcc uses hypoxanthine and 6-MP as sources
of nucleic acid purines, but SF/MP does not.
Moreover, SF/MPcc is distinguished from the wild
strain not only by its resistance to 6-MP but also
by the fact that it requires less folie acid for
optimal growth (17). It is quite possible, therefore,
that this mutant's resistance to 6-MP is due to
the high rate at which it can synthesize purines
de novo. In this connection it must be remembered
that 6-MP is not inhibitory to the wild strain when
sufficient exogenous purine is available, i.e., 6-MP
is inhibitory only if the supply of purines is limited.
Presumably in SF/MPcc the rate of purine syn
thesis de novo is so high that, under the conditions
of our experiments, the supply of purines was
always abundant. If the 6-MP resistance of SF/
MPcc is indeed due to the rapidity with which
it can synthesize purines de novo, it might be
anticipated that it should also be somewhat re
sistant to folie acid antagonists such as A-methopterin. This has been shown to be the case (17).
1 It has been shown that an enzyme from pigeon liver which
converts hypoxanthine and guanine to their nucleotides can
also synthesize a phosphoribosyl derivative of 6-MP from
6-MP (19).
3Work by Dr. R. W. Brockman (private communication)
has shown that SF/MP cannot convert these purines into their
free nucleotides, in confirmation of this hypothesis.
faecalis and 6-Mercaptopurine
443
The present experiments have disclosed no dif
ferences between the double mutants, SF/MP/A
and SF/A/MP, on the one hand, and the sensitive
mutants on the other. The varying ability of the
strains to use 6-MP as a purine source is almost
certainly a factor in 6-MP resistance. This is
particularly striking in the case of the double
mutant SF/A/MP and may provide an explana
tion for the resistance of this strain. There is a
precedent for the observation that a quantitative
change in the ability of bacteria to use a substrate
should result in qualitative changes in growth
response (1). In accordance with the mechanism
suggested above for the action of 6-MP (6), it is
possible that some of the postulated 6-MP nucleo
tide might be converted to inosinic acid and that
this might be sufficient to reverse the action of
the remaining 6-MP nucleotide.
It should perhaps be emphasized that, although
for convenience the present studies have been
limited to nucleic acids, it is not suggested that
6-MP exerts a specific effect on nucleic acid me
tabolism per se. On the contrary, it seems more
probable that it may inhibit conversion of inosinic
acid to adenine and guanine derivatives. If this
is so, its effects on tumors and microorganisms
may be due to its interfering with syntheses of
nucleic acids or purine-containing coenzymes or
both (7, 11, 12).
It seems not unlikely that the mechanisms of
the effects on tumors and microorganisms may
have much in common. It has been shown that
varying tissues including tumors depend upon de
novo synthesis to greater or lesser degrees relative
to their ability to use preformed purines (4, 9, 10).
Furthermore, two leukemias, Line I and Line I/A,
which differ in their sensitivity to A-methopterin,
differ also in the extent to which they utilize
exogenous formate for the synthesis of RNA pu
rines (2). If the view advanced above, that the
6-MP resistance of SF/MPcc is due to its increased
ability to synthesize purines de novo, is correct,
it is at least conceivable that the resistance of
certain tumors to antagonists of purine synthesis
or utilization may be explicable on similar lines.
SUMMARY
The effect of 6-mercaptopurine on S. faecalis
(ATCC No. 8043) and several of its mutants has
been investigated. Some mutants are able to use
the inhibitor as a source of nucleic acid purines;
to some extent, resistance may be due to that fact.
Two single mutants are resistant to 6-MP for other
reasons. The first appears to lack the enzyme
system which converts certain purines, including
6-MP, to nucleotides, and it is thought that the
Downloaded from cancerres.aacrjournals.org on June 11, 2017. © 1958 American Association for Cancer Research.
444
Cancer Research
nucleotide of 6-MP is the true inhibitor. The
second has a greater ability to synthesize purines
de novo and thus can better withstand the purine
antagonist. These facts emphasize the multiple
mechanisms which can lead to resistance.
ACKNOWLEDGMENTS
The authors wish to thank Dr. George Bosworth Brown
for his continued interest and very helpful discussions of this
project.
REFERENCES
1. BALIS,M. E.; BHOOKE,M. S.; BROWN,G. B.; and MAOASAMK, B. The Utilization of Purines by Purineless Mu
tants of Aerobacter acrogenes. 3. Biol. Chem., 219:917-26,
1956.
2. BALIS,M. E., and DANOIS,J. Effects of A-methopterin on
Nucleic Acid Synthesis in Leukemic Spleen Breis, Cancer
Research, 15:607-8, 1955.
3. BALIS,M. E., and HUTCHISON,D. J. The Utilization of 6Mercaptopurine by Streptococcusfaecalis. Fed. Proc., 16 :
149, 1957.
4. BALIS,M. E.; HTLIN, V.; COULTAS,M. K.; and HUTCHI
SON,D. J. Metabolism of Resistant Mutants of Streptococcu>faecalis. II. Incorporation of Exogenous Purines. Can
cer Research, 18:220-25, 1958.
5. BALIS,M. E., and LAKK,C. T. Synthesis of PNA and DNA
in Escherichia coli. Fed. Proc., 13:178, 1954.
6. BALIS,M. E.; LEVIN,D. H.; BROWN,G. B.; ELION,G. B.;
NATHAN,H. C.; and HITCHINOS,G. H. The Effects of 6Mercaptopurine on Lactobaclllus casei. Arch. Biochem. &
Biophys., 71:858-66, 1957.
7. BALIS,M. E.; LEVIN,D. H.; BROWN,G. B.; ELION,G. B.;
VANDERWERFF,
H.; and HITCHINQS,G. H. The Incorpora
tion of Exogenous Purines into Pentose Nucleic Acid by
Lactobacillus casei. J. Biol. Chem., 196:729-47, 1952.
8. BALIS,M. E.; LIVIN, D. H.; and LUZZATI,D. A PurineHistidine Relationship in Escherichia coli. J. Biol. Chem.,
216:9-16,1955.
Vol. 18, May, 1958
9. BALIS,M. E.; VANPRAAO,D.; and AEZEN,F. Studies on
the Metabolism of Human.Tumors. II. Pentosenucleic
Acid Synthesis in Tumor-bearing Rats. Cancer Research,
16:628-31, 1955.
10. BALIS,M. E.; VANPHAAO,D.; and BROWN,G. B. Studies
on the Metabolism of Human Tumors. I. Pentosenucleic
Acid Synthesis in Tumor-bearing Hamsters. Cancer Re
search, 16:673-78, 1955.
11. BOLTON,E. T., and MANDEL,H. G. The Effects of 6-Mercaptopurine on Biosynthesis in Escherichia coli. 3. Biol.
Chem.. 227:833-44, 1957.
12. BROWN,G. B., and BALIS,M. E. Nucleic Acids and Re
lated Derivatives as Targets for Chemotherapy. In The
Leukemias: Etiology, Pathophysiology and Treatment,
pp. 507-15. New York: Academic Press, Inc., 1957.
13. CLARKE,D. A.; PHILIPS,F. S.; STEHNBEHO,
S. S.; STOCK,
C. C.; ELION,G. B.; and HITCHINGS,G. H. 6-Mercaptopurine: Effects in Mouse Sarcoma 180 and in Normal Ani
mals. Cancer Research, 13:593-604, 1953.
14. ELION,G. B.; HITCHINOS,G. H.; and VANDERWERFF,
H.
Antagonists of Nucleic Acid Derivatives. VI. Purines.
J. Biol. Chem., 192:505-18, 1951.
15. ELION,G. B.; SINGER,S.; HITCHINGS,G. H.; BALIS,M. E.;
and BROWN,G. B. Effects of Purine Antagonists on a
Diaminopurine-Resistant Strain of Lactobacillus casei.
i. Biol. Chem., 202:647-54, 1953.
16. HUTCHISON,D. J. Biological Activities of 6-Mercaptopurine: Effects on Streptococcusfaecalis. Ann. N. Y. Acad.
Sc., 60:212-19, 1954.
17.
. Metabolism of Resistant Mutants of Streptococcus
faecalis. I. Isoktion and Characterization of the Mutants.
Cancer Research, 18:214-19, 1958.
18. LOBVINGEB,
R., and BERMAN,M. The Efficiency Criteria
in Radioactivity Counting. Nucleonics, 9:26-39, 1951.
19. LUKENS,L. N., and HEHHINGTON,
K. A. Enzymic Forma
tion of 6-Mercaptopurine Ribotide. Biochim. et Biophys.
Acta, 24:432-33, 1957.
20. SCHMIDT,G., and THANNHAUSER,
S. J. A Method for the
Determination of Deoxyribonucleic Acid, Ribonucleic
Acid and Phosphoproteins in Animal Tissues. J. Biol.
Chem., 161:83-89, 1945.
Downloaded from cancerres.aacrjournals.org on June 11, 2017. © 1958 American Association for Cancer Research.
Metabolism of Resistant Mutants of Streptococcus faecalis: III.
The Action of 6-Mercaptopurine
M. Earl Balis, Valia Hylin, M. Katharine Coultas, et al.
Cancer Res 1958;18:440-444.
Updated version
E-mail alerts
Reprints and
Subscriptions
Permissions
Access the most recent version of this article at:
http://cancerres.aacrjournals.org/content/18/4/440
Sign up to receive free email-alerts related to this article or journal.
To order reprints of this article or to subscribe to the journal, contact the AACR Publications
Department at [email protected].
To request permission to re-use all or part of this article, contact the AACR Publications
Department at [email protected].
Downloaded from cancerres.aacrjournals.org on June 11, 2017. © 1958 American Association for Cancer Research.