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Biosynthesis and regulation of superoxide

dismutase

Article in Free Radical Biology and Medicine · February 1988

DOI: 10.1016/0891-5849(88)90111-6 · Source: PubMed

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Free Radical Biology & Medicine. Vol. 5. pp. 377-385, 1988 0891-5849/88 $3.00+ .00
Printedin the USA. All rightsreserved. © 1988PergamonPressplc

Review Article
BIOSYNTHESIS AND REGULATION OF SUPEROXIDE DISMUTASES

HOSNI M. HASSAN
Departments of Food Science and Microbiology, North Carolina State University, Raleigh, NC 27695-7624, U.S.A.

Abstract--The past two decades have witnessed an explosion in our understanding of oxygen toxicity. The
discovery of superoxide dismutases (SODs) (EC. 1.15.1.1), which specifically catalyze the dismutation of super-
oxide radicals (02-) to hydrogen peroxide (H202) and oxygen, has indicated that O2- is a normal and common
byproduct of oxygen metabolism. There is an increasing evidence to support the conclusion that superoxide radicals
play a major role in cellular injury, mutagenesis, and many diseases. In all cases SODs have been shown to protect
the cells against these deleterious effects. Recent advances in molecular biology and the isolation of different SOD
genes and SOD c-DNAs have been useful in. proving beyond doubt the physiological function of the enzyme. The
biosynthesis of SODs, in most biological systems, is under rigorous controls. In general, exposure to increased
pO2, increased intracellular fluxes of O2-, metal ions perturbation, and exposures to several environmental oxidants
have been shown to influence the rate of SOD synthesis in both prokaryotic and eukaryotic organisms. Recent
developments in the mechanism of regulation of the manganese-containing superoxide dismutase of Escherichia
coli will certainly open new research avenues to better understand the regulation of SODs in other organisms.

Keywords--pO2, O2-, Growth rate, Metal depletion/repletion, Anaerobic synthesis of MnSOD, Redox-sensitive
repressor, Autogenous regulation, Environmental stresses, Free radical

INTRODUCTION and .OH) generated during the normal biological re-

Superoxide dismutases (SODs) (EC. 1.15.1.1) are me-
talloenzymes that are widely distributed among oxy-
gen-consuming organisms, aerotolerant anaerobes, and
some obligate anaerobes. They are essential for the
defense against oxygen toxicity, which is mediated by
the partially reduced oxygen intermediates (O2-, HzO2,
Paper number 11454 of the Journal Series of the North Carolina
Agricultural Research Service, Raleigh, NC 27695. The use of trade
names in this publication does not imply endorsement by the North
Carolina Agricultural Research Service of the products named, nor
criticism of similar ones not mentioned.
Hosni M. Hassan was born in 1937 in Alexandria, Egypt. He
received his B. Sc (Agriculture) in 1959 from the University of Ain-
Shams, Cairo; and his Ph.D. (Microbiology)in 1967, from the Uni-
versity of California at Davis (UCD). He was an assistant professor
of microbiology and biochemistry at Cairo High Polytechnical In-
stitute, then at the University of Alexandria. In 1972, he moved
back to North America. He was a visiting professor at the Macdonald
College of McGill University, Assistant Microbiologist at the Uni-
versity of Maine, and a Research Associate in Fridovich's laboratory
at Duke University. Currently he is a Professor of Food Science,
Microbiologyand Toxicologyat N.C. State University. Dr. Hassan's
research interests are in the areas of Microbial Physiology and En-
zymology. During the past 13 years, he has been interested in oxygen
toxicity, biological roles of antioxidant enzymes, and the biosyn-
thesis and regulation of superoxide dismutases in microorganisms.
Dr. Hassan is an American Fulbright Senior Research-Scholar in
France. 1987-1988. He enjoys travel, photography, walking, and
reading.
377
duction of dioxygen. SODs are unique in that their
substrate is a free radical and that they catalyze the
dismutation of O2- to H20: and 02 at a very fast rate,
2 x 109 M-Is -1.
SODs, isolated from a wide range of organisms, fall
into three types depending on the metal found in their
active center. McCord and Fridovich ~were the first to
describe the superoxide-dismuting ability of a green
copper-containing protein that was isolated some 30
years earlier,: and thought to be a copper storage pro-
tein. This enzyme is now known to contain both copper
and zinc (CuZnSOD) where the zinc plays a structural
role. The other two types of SODs were soon discov-
ered: one contains manganese (MnSOD)) and the sec-
ond contains iron (FeSOD)? The distribution of these
three forms of SODs is distinctly different, and may
provide an interesting evolutionary scheme. The
CuZnSODs are typically found in the cytosol of eu-
karyotes, while FeSODs are found in prokaryotes. On
the other hand, MnSODs are found in prokaryotes and
in mitochondria. There are, however, some exceptions
to this simplified evolutionary scheme (For a recent
complete account, see Bannister et al)). Amino acid
sequence data show that the three types of SODs fall
into two distinct phylogenetic families, the CuZnSOD
378 H.M. HASSAN
and the Fe/MnSOD. There is no sequence homology
or secondary structure similarity between the two fam-
ilies. 6 The FeSODs and MnSODs show a high degree
of amino acid sequence and structural homologies, while
they are completely unrelated to CuZnSODs. 7.s Thus,
it is clear that these two families of SODs have evolved
independently in response to a common selective pres-
sure, that is, the evolution of oxygen and the threat of
its toxicity. In this review I will concentrate on the
biological function and the biosynthesis of SODs in
response to various environmental stimuli or stresses
in an attempt to understand the regulatory mecha-
nism(s) that govern their biosynthesis in different liv-
ing cells.
BIOLOGICAL FUNCTION
All living cells are prone to oxygen toxicity, 9-~1 and
mutagenicity. 12-16Several lines of evidence are avail-
able which support the superoxide theory of oxygen
toxicity and the protective role of superoxide dismu-
tases. 17.LsMcCord et al. 19 reported that aerobic organ-
isms contain more SOD than aerotolerant organisms,
while anaerobes have no detectable SOD activity. Later
studies demonstrated the presence of SOD in some
obligate anaerobes, 2°-22 and a positive correlation is
found between the degree of oxygen tolerance and the
level of SOD in anaerobic organisms. 23 The presence
of low concentrations of SOD in many anaerobes may
represent an evolutionary response to ensure their sur-
vival during transient exposures to oxygen which.they
encounter in route from one anaerobic niche to another.
The finding that some anaerobes are able to induce
SOD when exposed to very low concentrations of ox-
ygen, 24 lends further support to the biological role of
SOD as an antioxidant enzyme.
Increased levels of MnSOD in E. coli, induced by
a variety of physiological conditions, impart resistance
against oxygen toxicity, 25-33 and against the mutagen-
icity of oxyradicals. 15.34The physiological role of SODs
in protecting E. coli against the toxicity and mutagen-
icity of oxyradicals has been further explored in mu-
tants lacking MnSOD (sodA), FeSOD (sodB), or
both.16'35 The double mutant (sodA-sodB) is unable to
grow aerobically in minimal medium, but grows nor-
mally in absence of oxygen. In rich medium, the sodB
mutant exhibits the same sensitivity towards paraquat,
an oxyradical generator, as the wild-type strain, whereas
the soda mutant is more sensitive to paraquat than the
sodB mutant or the wild-type. In minimal medium,
however, both soda and sodB mutants are more sen-
sitive to paraquat than the wild-type.3~ The introduction
of a plasmid overproducing Mn- or FeSOD into the
double mutant restores its wild type phenotype with
regard to aerobic growth on minimal medium and tol-
erance to paraquat. 35 The oxygen-dependent enhance-
ment of mutagenesis in the sod-negative mutants is
independent of RecA but dependent on the presence of
exonuclease III.16 Oxygen is more mutagenic in sodA-
sodB and sodA mutants than in sodB mutant or in the
wild-type strain. ~6Conversely, the presence of a plas-
mid overproducing either FeSOD or MnSOD reduces
the mutagenic frequency seen in the double mutant
(sodA-sodB) to that of the wild-type.
The recent advances in molecular biology and the
isolation of SOD genes and SOD c-DNAs have facil-
itated more direct studies into the physiological func-
tion of superoxide dismutases. 36 Thus, the human
copper-zinc superoxide dismutase (h-CuZnSOD) com-
plements SOD-deficiency in E. c0li.37 The double mu-
tant (sodA-sodB) is unable to grow aerobically in min-
imal medium and growth in rich medium is inhibited
by the presence of paraquat or hydrogen peroxide,
however, the expression of the human enzyme in this
mutant converts it back to the wild-type phenotype.
When the human-CuZnSOD gene is transfected into
human Hela cells or mouse L-cells, transformants ex-
pressing elevated levels of h-CuZnSOD are more re-
sistant to paraquat than normal cells/8 Similar results
are obtained with cultured N IH/3T3 mammalian cells
transfected with a cDNA expressing high levels of the
human-CuZnSOD. 39Furthermore, SOD is essential for
the activity of E. coli ribonucleotide reductase. ~°4~
Recent studies have demonstrated the direct toxicity
of 02- without invoking the metal-catalyzed Haber-
Weiss reaction. For example: (a) in E. coli, a,13-di-
hydroxyisovalerate dehydrogenase, an enzyme essen-
tial for the biosynthesis of branch-chain amino acids,
is extremely sensitive to increasing intracellular fluxes
of 02- caused by the presence of redox-active com-
pounds in the growth medium; 42 (b) bovine liver cat-
alase is inactivated by O2- and SOD protects; 43 and
(c) exposure of a purified glutathione peroxidase to an
enzymatic source of O2- plus H202 causes inactivation
of the enzyme which is preventable by SOD, but not
by catalase. .4
These findings clearly support the conclusion that
superoxide radicals are toxic or may yield toxic prod-
ucts, and that superoxide dismutases provide, in vivo,
cellular protection by virtue of their ability to catalyt-
ically dismute O2-. 17,18,45
BIOSYNTHESIS AND REGULATION
Exposure of living cells to physiological extremes
is known to evoke several global cellular responses
designed to provide cellular protection and to insure
the survival of the species. The finding that superoxide
 Control of SOD biosynthesis
dismutases are important in normal aerobic survival
aroused the interest of many biologists, biochemists,
and geneticists, to attempt to understand the regulation
of SODs in different organisms. The findings clearly
demonstrate that the biosynthesis of superoxide dis-
mutases, in most biological systems, are under rigorous
controls.
Prokaryotes
Prokaryotes exhibit the broadest range of diversity
with regard to their oxygen requirement and tolerance,
which makes them an ideal model system for studying
the regulation of SODs biosynthesis. Prokaryotes are
also unique in their ability to grow fast and to readjust
their cellular composition to meet the fast and sudden
changes in their environment. Cellular modulation, ac-
complished via induction, repression, derepression,
a c t i v a t i o n . . , etc., is the key for achieving fast ad-
aptation and the best cellular economy. Indeed, mod-
ulation of SOD biosynthesis in a given organism is
finely-tuned to protect the cells against oxidative
damages.
Synthesis of SOD in response to changes in p02. Aer-
obically grown E. coli contains three distinct isozymes
of superoxide dismutase,~ one of which contains man-
ganese, 3 the second contains iron, 4 and the third is a
hybrid consisting of one subunit from each of the other
two isozymes and originally thought to contain only
iron. 47 More recent biochemical48 and radiolabeling49
studies have clearly demonstrated that the hybrid en-
zyme contains both iron and manganese. Anaerobically
grown E. coli, in contrast, contains only FeSOD. ~
Thus, the iron-enzyme is present under both anaerobic
and aerobic conditions and is, therefore, constitutive.
In contrast the Mn- and the hybrid-forms are absent
under anaerobic conditions but are rapidly synthesized
upon exposure to air. 46 It has been proposed that the
presence of a constitutive FeSOD in E. coli provides
a back-up defense mechanism against sudden aerobic
exposures to oxygen) ° The anaerobe, Bacteroidesfra-
gilis contains low levels of FeSOD when grown ana-
erobically, and exposure to an atmosphere containing
2% oxygen induces the enzyme. 24 Exposure to high
concentrations of oxygen have been shown to induce
MnSOD in E. coli, 5~ Streptococcus faecalis,51's2 Pro-
teus vulgaris, s2 Enterobacter cloacae,~2 Staphylococ-
cus aureus, 52 Staphylococcus epidermidis, 52 Listeria
monocytogenes,S2 and Streptococcus sanguis,Sa as well
as FeSOD in Rhizobium japonicum, ~ Bdellovibrio
stolpii, 55 Alcaligenes faecalis, 52 and Pseudomonas
aeruginosa) 2 P. vulgaris, E. cloacae, Salmonella ty-
phimurium, and E. coli all belong to the Enterobac-
379
teriaceae family and all possess a constitutive FeSOD
and an inducible MnSOD. 52 It is clear that the mech-
anisms for biosynthesis and regulation of SODs in pro-
karyotes are not universal. Therefore, in the following
sections I will concentrate on the biosynthesis and reg-
ulation of MnSOD with special reference to E. coli.
Synthesis of SOD in response to changes in specific
growth rate and metabolism. Growth of E. coli in a
glucose-limited chemostat culture maintained under
constant and abundant aeration demonstrated that the
rate of MnSOD biosynthesis is proportional to the spe-
cific rate of growth and the rate of respiration. 2s These
results indicate that the inducer for MnSOD cannot be
molecular oxygen, per se, but rather a unique product
of its metabolism. These results also advanced the hy-
pothesis that the concentration of MnSOD in the cells
is modulated to correlate with the intracellular flux of
O2- generated during aerobic growth. It was also noted
that when E. coli is grown in rich medium containing
limited amounts of glucose (i.e., trypticase-soy/yeast
extract, TSY) the cells' content of MnSOD remains
low while utilizing glucose but increases after glucose
is exhausted. 26 In general, if E. coli is utilizing a fer-
mentable carbon source, such as glucose, the MnSOD
level is much lower than when an oxidizable carbon
source, such as succinate, lactate, or trypticase-pep-
tone, is utilized. 2~The lower MnSOD level seen when
the cells are utilizing glucose is not due to catabolite
repression. ~ These results, led to the conclusion that
the rate of intracellular flux of 02- in E. coli is low
during glucose fermentation, but increases during ox-
idative metabolism and that the cells are capable of
modulating their content of MnSOD to meet their
changing needs for the scavenging of 02-. This sup-
position led us to study the effects of redox-cycling
compounds, capable of artificially increasing the in-
tracellular flux of O2-, on the rate of MnSOD biosyn-
thesis in E. coli. 26"27"56
Synthesis of SOD in response to increased intracel-
lular/ extracellular flaxes of 02-. Several redox-cy-
cling compounds can enter E. coli, 27'56 and other
bacterias2 and cause increase in the intracellular pro-
duction of O2-. In general, these compounds are first
reduced by nonspecific diaphorases present in the cells;
the reduced forms are rapidly reoxidized by oxygen to
generate 02-.s6 The generation of 02- by these redox-
active compounds is dependent upon their ability to
enter the cells, the presence of oxygen, and on the
presence of a metabolizable substrate that can provide
a reducing equivalent. 28 Paraquat, pyocyanine, strep-
tonigrin, juglone, plumbagin, menadione, methylene
blue, and azure C are among the compounds which
 380 H.M. I-IASSAN
function in this way. 56 They all act catalytically to
increase the intracellular flux of O2- and in doing so
they increase the rate of cyanide-insensitive respira-
tion, 27"56since they provide a cytochrome-independent
pathway to oxygen. These compounds exhibit an ox-
ygen-dependent toxicity, and in a real sense they ex-
acerbate the toxicity of o x y g e n : 7 The cells respond to
such threat by inducing MnSOD. Kinetic analysis of
MnSOD induction by paraquat, showed that the ad-
dition of 0.5 mM paraquat to aerobically growing cul-
ture results, after a brief lag, in a rapid linear increase
in MnSOD. 27 Removal of paraquat from the growth
medium causes a sharp decline in the rate of MnSOD
biosynthesis. The induction of MnSOD by paraquat is
prevented by inhibitors of transcription or translation,
but not by inhibitors of replication. 56 Paraquat also
induces MnSOD in P. vulgaris, 52 E. cloacae: 2 S. au-
reus, 52 and S. typhimurium. ~5Similar inductive effects
are seen with all the redox-active compounds listed
above, s6 On the other hand, extracellularly generated
0 2 - , via the action of xanthine/xanthine oxidase or
riboflavin and light, does not induce MnSOD in E.
coli, due to the fact the O2- does not cross the E. coli
cell envelope. 3t These results indicate that the induc-
tion of MnSOD in E. coli is under transcriptional con-
trol and that intracellular 02- or a product thereof is
the effector.
Synthesis of SOD in response to metal depletion~re-
pletion. The role of iron in the regulation of MnSOD
biosynthesis in E. coli was accidentally discovered dur-
ing a study on the effects of inhibitors of DNA bio-
synthesis on the induction of MnSOD by paraquat. A
2-fold increase in MnSOD is noted when nalidixic acid
is used, instead of hydroxyurea, as an inhibitor of DNA
biosynthesis) 8 Nalidixic acid has a synergistic effect
on the induction of MnSOD by paraquat (Hassan, un-
published data). It has been shown that the induction
of MnSOD by nalidixic acid is not due to its known
ability to inhibit DNA gyrase nor to increase the in-
tracellular flux of O2-, but rather due to its ability to
chelate ferrous i r o n ) 8 Further studies have also shown
that the addition of excess iron to the growth media of
E. coli has a repressive effect on the biosynthesis of
MnSOD under normal (aerobic) or inducing conditions
(i.e., in presence of 0.2 mM paraquat)) 9'6° Addition
of manganese to cultures already derepressed for MnSOD
biosynthesis causes a further increase in the amount of
active enzyme, 6° probably due to a post-translational
activation of the Mn-apoenzyme. 49 Removal of metals
from the growth medium by Chelex-lO0 also de-
represses the synthesis of MnSOD, but repletion of
the medium with iron abolishes this e f f e c t : ° Further-
more, chelators specific for ferrous iron, that is,
2,2'-dipyridyl, 1,10-phenanthroline, 8-OH-quinoline.
and ferrocyphen, cause a 3- to 7-fold increase in
M n S O D ) 8-6° The iron chelator, 2,2'-dipyridyl, also
induces MnSOD in P. vulgaris, E. cloacae, S. aureus.
and S. typhimurium: 2 Most fascinating, however, is
the finding that ferrous iron chelators can also induce
MnSOD in E. coli in the absence of dioxygen. 6° A
model has been proposed which accommodates all of
the above findings. 6° In this model, the synthesis of
MnSOD is envisioned as being regulated by a negative-
control operon, where the repressor protein (RP) con-
tains iron (Fig. 1). It is proposed that the active re-
pressor contains ferrous iron (RP-Fe-'+), while the
inactive form contains either ferric iron (RP-Fe 3+) or
no iron (RP). Accordingly, in the absence of oxygen
the active repressor will be predominant and no MnSOD
is made. On the other hand, conditions known to ox-
idize Fe 2+ to Fe 3+ (i.e., oxygen or oxyradicals) or to
remove Fe 2+ from the cells (i.e., iron chelators) will
generate the inactive forms of the repressor and will
result in the synthesis of the enzyme, even in the ab-
sence of oxygen. This model also suggests that con-
ditions known to change the redox status of the cell
may also affect the synthesis of MnSOD in absence of
oxygen. Indeed, this is the case. 49'6t Thus, under an-
aerobic conditions, active MnSOD is induced in the
presence of potassium ferricyanide, copper-cyanide
complex, ammonium persulfate, and hydrogen per-
oxide, 6~ as well as in the presence of nitrate plus para-
quat. 49 Western blot analysis revealed that the induc-
tion of MnSOD by these oxidants is associated with
de novo protein biosynthesis. 61 Furthermore, the pres-
ence of increasing concentrations of ozone, a powerful
oxidant, induces MnSOD in E. coli. 62 These results
suggest that the redox state of the cell influences the
biosynthesis and/or the activity of the proposed iron-
I to I I PI~ o I
SG
jl
/ MnSOD
RP" RP-Fe 3+ I
's
Fe2+ O~ ,,/
-o~,,'
ll~.Fe 2'"
Fig. 1. Schematicmodel for the regulation of MnSOD by oxygen,
superoxideradicals, and ferrousiron chelators. RG, regulatorygene;
P, promotor; O, operator; SG, structural gene for MnSOD; RP.
aporepressorprotein (inactive);RP-Fe3*, ferric repressor (inactive);
RP-Fe2*, ferrous repressor (active). (From Moody, C. S.; Hassan,
H. M., J. Biol. Chem 259:12821-12825; 1984. Reprinted by per-
mission.)
 Control of SOD biosynthesis
containing repressor that serves to negatively regulate
MnSOD biosynthesis. The utilization of reduction po-
tential in the regulation of MnSOD biosynthesis in E.
coli seems logical when viewed in the following light.
Facultative anaerobes take advantage of both aero-
biosis and anaerobiosis. Exposure to oxygen, however,
poses problems arising from oxygen toxicity. E. coil
overcomes oxygen toxicity by inducing antioxidant en-
zymes, MnSOD in particular. The reduction potential
is a very sensitive but complex indicator of the inte-
grated physiological state of the cell. Thus, very small
changes in oxygen concentration can result in large
changes in reduction potential. If E. coil is pro-
grammed to respond to changes in reduction potential
instead of oxygen concentrations, then the cells could
induce the antioxidant enzymes at very low pO2 and
thus be poised and ready to combat the toxic effects
of oxygen when confronted with a higher pO:. 6~
Another model has been proposed to account for the
effects of iron, manganese and chelators on the bio-
synthesis of both Mn- and Fe-SODs in E. c o l l . 63-65 In
this model (Fig. 2), both FeSOD and MnSOD are viewed
as autogenously regulated catalysts whose conversion
from inactive apo-proteins to catalytically active holo-
enzymes depends on the availability of the appropriate
metal cofactor and upon a prerequisit unknown con-
centration of O:- capable of converting sufficient amount
of Mn2÷ to Mn 3+ in order to compete effectively with
Aulooenous
Fe-m-opo (inactive)
AMINO ACIDS
~~. ~.~FMn-f-OlDO(Inactive)
Ao;.,,.72," F. lA,,,,.i
Mn(I~),O~ . 2 H ° " M*llllloH20- /
Fe(lll).O~ . • Fe{ll)*O 2
Fig. 2. Effects of metals on the biosynthesis of SODs--a proposal.
Abbreviations: m-apo, apoprotein of MnSOD; f-apo, apoprotein of
FeSOD. The valences of the metals are not shown. It is assumed
that Fe(lI) binds to both m-apo mad f-apo more avidly than Mn(lI),
but not more avidly than Mn(III). The reactions illustrated at the
bottom indicate why O:- increases the availability of Mn(III). (From
Pugh, S. Y.; Fridovich, I. J. Bacteriol. 162:196-202; 1985. Re-
printed by permission.)
381
Fe2+ for the active sites in the apo-proteins. Thus,
under anaerobic conditions the apo-MnSOD is sup-
posed to be made but the active site is occupied by
iron and is therefore catalytically inactive. Aerobically
and in presence of O2--generators, Mn2+ is supposed
to be oxidized to Mn 3+ which in-turn is a strong com-
petitor for the active site of apo-MnSOD and therefore
generates catalytically active MnSOD. 65 This model
also proposes autogenous repression by the apo-pro-
teins and by the inactive enzymes that contain the in-
correct metals.~'65 Fortunately, both regulatory models
are testable.
Recent studies, 49 have demonstrated that 59Fe binds
to all the dismutase isozymes of E. coli, whereas 54Mn
is more specific for the manganese and the hybrid iso-
zymes. In the presence of paraquat, an Oe- generator,
more 59Fe is incorporated into MnSOD, whereas no
54Mn is incorporated into FeSOD. 49These data are the
opposite to what is predicted by the autoregulatory
model.65 Furthermore, the inactive form of the MnSOD
(i.e., the one containing 59Fe) appears to have no au-
toregulatory function, since its abundant presence does
not prevent the induction of MnSOD by paraquat. 49
Preliminary evidence was also presented for the pres-
ence of two unique iron-proteins49that may qualify for
the repressor function proposed in the transcriptional
repression model. 6° Furthermore, the presence of a
multicopy plasmid carrying the MnSOD gene (SodA)
was shown to cause the anaerobic expression of the
gene, presumably by neutralizing the limited number
of repressor molecules found in the cells. 49'67The most
convincing arguments in support of the negatively con-
trolled operon, 6° come from the following findings: (a)
western blot analysis of cell-free extracts prepared from
anaerobically grown cells shows the absence of MnSOD-
antigen;35"6~ (b) the DNA sequence of sodA shows a
region of dyad symmetry at the - 35 region which may
serve as a binding site for a repressor molecule;~ (c)
13-galactosidase is induced by paraquat in a strain of
E. coil carrying sodA-lacZ fusion;67 (d) MnSOD is
induced anaerobically by IPTG in ptac-sodA operon
fusion strains;68 and (e) MnSOD is induced by oxygen
strains overproducing FeSOD. 69
These results clearly indicate: (a) the absence of
active or inactive MnSOD in anaerobically grown cells;
(b) the gene for MnSOD possesses a potential binding
site for a regulatory protein; (c) the induction of the
soda gene by paraquat is controlled at the transcrip-
tional level and manganese ions play no regulatory
role; (d) under anaerobic conditions, manganese ions
of the proper valence are present in high concentration
to allow the synthesis of about 60 U of active MnSOD
per mg of protein, as seen in the ptac-sodA operon
fusion strains; and (e) superoxide radicals are not di-
382 H.M. HASSAN
rectly involved in the regulation of MnSOD. Further-
more, MnSOD regulatory mutants have recently been
isolated (Hassan and Touati, manuscript in prepara-
tion) and found to express the soda gene anaerobically
and to be uninducible by paraquat or chelators. In toto,
the data support the proposed modeP ° for a negatively
controlled operon where the repressor molecule is now
envisioned as an allosteric redox sensing protein. 61
However, the recent work by TouatP s suggests the
presence of a positive transcriptional control by O2-,
and an autogenous control that does not require the
whole apoprotein, besides the originally proposed 6°
negative transcriptional control by an iron-repressor.
Indeed, the availability of regulatory mutants will al-
low a better understanding of MnSOD regulation.
Effect of metal substitution on the activity and synthesis
of SODs. Metal substitution studies are valuable tools
in understanding the structure/function of SODs and
the catalytic role of the native metal. Removal of the
metal from SODs results in loss of activity. Attempts
at metal-replacements with FeSODs and MnSODs from
different bacterial species have demonstrated strict metal
cofactor specificity. 7° Earlier studies with Fe/Mn-
SODs, from E. coli, have demonstrated that substi-
tution of iron for manganese, or vice versa, results in
inactive enzymes. 71,7z Thus, it is clear that the two
enzymes, in spite of their relatedness, 6'8 must have
diverged in a way that only the native metals can restore
their catalytic activity.
The story is different, however, for the FeSODs
from anaerobic organisms, where the protein can be
reconstituted or modulated during biosynthesis to yield
an active enzyme containing manganese in place of
iron. 73-76 Thus, the FeSOD from B. fragilis can be
reconstituted as a Mn-containing enzyme o73 Further-
more, anaerobically grown B. fragilis contains FeSOD,
but oxygen-stressed cells apparently use the same apo-
protein to make an active MnSOD. 74 Also, Propioni-
bacterium shermanii, 75 and Streptococcus mutans, 76
are able to synthesize either FeSOD or MnSOD, using
the same apoprotein, depending on the metal supplied
in their growth media. The evolutionary significance
of this, cambialistic, 76 class of enzymes is not clear.
However, it is interesting to note that they are from
cells that do not use oxygen as an electron-acceptor,
and only possess the FeSOD gene. Accordingly, one
may propose that organisms capable of using oxygen
as a terminal electron-acceptor or possess MnSOD gene
or both Fe/MnSOD genes may not be able to exchange
their metal-cofactors and remain active. This possi-
bility needs to be tested. Also, complete amino acid
sequences and X-ray crystallographic data for these
"cambialistic" enzymes are needed in order to be able
to compare their ligand fields with the more fastidious
Fe/MnSODs.
Synthesis of SOD in response to other environmental
stresses. Global cellular responses designed to provide
protection against several environmental stress factors
have been described. 77-79For example, the S O S , 77 heat
shock, 78 and oxidative s t r e s s 79 responses are induced
by DNA damaging agents, exposure to high temper-
atures, and exposure to hydrogen peroxide; and are
controlled by the recA, htpR, and oxyR gene products,
respectively, Oxyradicals have been shown to cause
DNA damage; s° and to induce the SOS response, 81
endonuclease IV, s2 and a new type of DNA repair
pathway. 83 The possibility that the synthesis of
MnSOD is under the control of these global stress
regulons has been tested by several investigators. Thus,
the regulation of the MnSOD in E. coli has been shown
to be independent of the inducible DNA repair (SOS)
system. 84On the other hand, oxyR mutants were shown
to contain more MnSOD, 79 and exposure of E. coli
culture to 48°C induced MnSOD upon return of the
culture to 37°C. 8~ However, recent studies on the ef-
fects of paraquat on the synthesis of MnSOD (Lee and
Hassan, and Bowen and Hassan, manuscripts in prep-
aration) and on the synthesis of 13-galactosidase using
an sodA-lacZ fusion, 69 in mutants deficient in heat
shock response (htpR) or in oxidative stress response
(oxyR), have clearly demonstrated that the regulation
of MnSOD is independent of these two regulons. The
possibility of a superoxide inducible (soi) regulon is
being examined by Kogema et al. 86
Genetic studies. The genes for MnSOD (sodA) and
FeSOD (sodB) of E. coli have been cloned, 87'88 and
sequenced. 66"89The DNA sequence for sodA indicates
that the gene constitutes a single operon having two
possible promoters, but only one of them appears active
under normal aerobic conditions. 66 The function of the
second promoter is not known at the present time, but
its involvement under highly inducing conditions is a
viable possibility. As discussed above, the sequence
also shows the presence of an almost perfect 19-base
palindrome at the - 3 5 region which represents a po-
tential binding site for a regulatory molecule. 66 The
linearized sodA is a better substrate for in vitro tran-
scription than the supercoiled f o r m , 66 this may be re-
lated, in part, to our earlier observation on the effect
of nalidixic acid on MnSOD biosynthesis. 58 On the
other hand, the DNA sequence of the sodB gene shows
the lack of a regulatory element, 89 which is in agree-
ment with the fact that FeSOD, in E. coli, is a con-
stitutive enzyme. 46 For a complete review the reader
is referred to the chapter by Touati. 36
 Control of SOD biosynthesis
Eukaryotes
The biosynthesis of SODs in eukaryotes in response
to increased oxygen concentrations, the presence of
superoxide radical generators, and the availability of
metals is not much different from that reported in
prokaryotes.
S a c c h a r o m y c e s cerevisiae has been used as a model
system to study the biosynthesis of SODs in eukar-
yotes. Under aerobic conditions, a cytochrome-c de-
ficient mutant is found to make twice as much SOD
as its isogenic wild-type strain. 9° The increased syn-
thesis of SOD in the mutant is due to increased intra-
cellular flux of O2- caused by the lack of a functional
electron transport chain, as depicted from the increased
rate of cyanide-insensitive respiration. 9° Addition of
paraquat also causes an increase in the rate of cyanide-
insensitive respiration and induces the synthesis of
MnSOD and CuZnSOD; the addition of copper further
stimulates the synthesis of CuZnSOD. 9~ Similar find-
ings were reported for S. cerevisiae growing in che-
mostat cultures. 92 Feeding chicks a diet low in copper
(<1 m g / k g dry matter) results in a significant decrease
in the activity of erythrocytes' SOD compared to that
of copper-supplemented (10 m g / k g dry matter) con-
trois.93 Subcutaneous injection of copper sulfate (5 rag/
kg body weight) in rats causes an increase in the ac-
tivity of CuZnSODs found in the cytosol and the mi-
tochondrial inter-membrane spaces. 94 The effect of
copper on SOD biosynthesis was also observed in the
fungus D a c t y l i u m dendroides, which contains both
CuZnSOD and MnSOD. 95 The fungus makes less
CuZnSOD but more MnSOD, when grown in a copper-
deficient medium, so that the total SOD in the cell
remains unaffected by the copper deficiency.
The synthesis of SOD has been shown to increase
in response to increased pO2 in yeast, 96 rat lung, 97,9s
endothelial cells, 99 and in maize leaves. ~00The air po-
lutant SO2 was shown to induce SOD in the leaves of
poplar tree, ~°~ but not in maize leaves. ~°° Exposure of
ten-day-old maize leaves to 10 -5 M paraquat for twelve
hours results in 40% increase in total SOD activity. ~02
More significant increases are seen in chloroplast and
cytosolic SODs (CuZnSODs) than in the mitochondrial
MnSOD. The increases in cytosolic and mitochondrial
SODs are the result of increased synthesis of these
proteins. As in prokaryotes, heat-shock treatment does
not increase the biosynthesis of SOD in maize.~°3 For
a complete review on the molecular biology of eukar-
yotic superoxide dismutases see Touati. 36
Acknowledgements--This work was supported by DMB-8609239
from the National Science Foundation. H.M.H. is an American Ful-
bright Senior Research-Scholar at the Institute of Jacques Monod,
University of Paris 7, Paris, France.
383
REFERENCES
1. McCord, J. M.; Fridovich, I. Superoxide dismutase: an en-
zymatic function for erythrocuprein (bemocuprein). J. Biol.
Chem. 244:6049-6055; 1969.
2. Mann, T.; Keilin, D. Hvmocuprein and Hvpatocuprein, cop-
per-protein compounds of blood and liver in mammals. Proc.
R. Soc. London B. Biol. Sci. 126:303-315; 1938.
3. Keele, B. B.; McCord, J. M.; Fridovich, I. Superoxide dis-
mutase from Escherichia coli B: A new manganese-containing
enzyme. J. Biol. Chem. 245:6176-6181; 1970.
4. Yost, F. J.; Fridovich, I. An iron-containing superoxide dis-
mutase from Escherichia call B. J. Biol. Chem. 248:4905-
4908; 1973.
5. Bannister, J. V.; Bannister, W. H.; Rotilio, G. Aspects of the
structure, function, and applications of superoxide dismutase.
C R C CriticalRev. Biochem. 22:111-180; 1987.
6. Harris, J. I.; Auffret, A. D.; Northrop, F. D.; Walker, J. E.
Structural comparisons of superoxide dismutase. Eur. J.
Biochem. 106:297-303; 1980.
7. Steinman, H. M.; Hill, R. L. Sequence homologies among
bacterialand mitochondrial superoxide dismutases. Proc. Natl.
Acad. Sci. USA 70:3725-3729; 1973.
8. Stallings,W. C.; Pauridge, K. A.; Strong, R. K.; Ludwig, M.
L. Manganese and iron superoxide dismutase are structural
homologs. J. Biol. Chem. 259:10695-10699; 1984.
9. Haugaard, N. Cellular mechanisms of oxygen toxicity.Phys-
iol. Rev. 48:311-373; 1968.
I0. Gottleib, S. E Effects of hyperbaric oxygen on microorga-
nisms. Annu. Rev. Microbiol. 25:I 11-152; 1971.
I I. Fridovich. I. The biology of oxygen radicals.Science 201:875-
880; 1978.
12. Fvnn, W. O.; Gerschman, R.~ Gilbert, D. L.; Terwilliger, D.
E.; Gothran, F. V. Mutagvnic effects of high oxygen tensions
on Escherichia coil Proc. Natl. Acad. 5ci. USA 43:1027-
1032; 1958.
13. Grifford, G. D. Mutation of an auxo~ophic strain of Esche~
richia coli by high pressure oxygen. Biochem. Biophys. Res.
Commun. 33:294-298; 1968.
14. Bruyninckx, W. J.; Mason, H. S.; Morse, S. A. Are physio-
logical oxygen concentrations mutagenic? Nature (London)
274:606-607; 1978.
15. Moody, C. S.; Hassan, H. M. Mutagenicity of oxygen free
radicals. Proc. Natl. Acad. Sci. USA 79:2855-2859; 1982.
16. Farr, S. B.; D'Ari, R.; Touati, D. Oxygen-dependent muta-
genesis in Escherichia coil lacking superoxidedismutase. Proc.
Natl. Acad. Sci. USA 83:8268-8272; 1986.
17. Fridovich, I. Superoxide dismutases. Annu. Rev. Biochem.
44:147-159; 1975.
18. Fridovich. I. Superoxide dismutases. Adv. Enzymol. 58:62-
97; 1986.
19. McCord, J. M.; Keele, B. B.; Fridovich, I. An enzyme-based
theory of obligate anaerobiosis: the physiological function of
superoxide dismutase. Proc. Natl. Acad. Sci. USA 68:1024-
1027; 1971.
20. Hewitt, J.; Morris, J. G. Superoxide dismutase in some
obligately anaerobic bacteria. FEBS Left. $0:315-318;
1975.
21. Carlsson, J.; Wrethen, J.; Beckman, G. Superoxide dismutase
in Bacteriodes fragilis and relatedBacteroides species.J. Clin.
Microbiol. 6:280-284; 1977.
22. Kirby, T. W.; Lancaster, J. R.; Fridovich, I. Isolation and
characterization of the iron-containing superoxide dismutase
of Methanobacterium bryantii. Arch. Biochem. Biophys.
210:140-148; 1981.
23. Tally, F. P.; Goldin, B. R.; Jacobus, N. B.; Gorbach, S. L.
Superoxide dismutase in anaerobic bacteria of clinical signif-
icance. Infect. lmmun. 16:20-25; 1977.
24. Privalle, C. T.; Gregory, E. M. Superoxide dismutase and O:
lethality in Bacteroides fragilis. J. Bacteriol. 138:139-145;
1979.
25. Hassan, H. M.; Fridovich, I. Physiological function of super-
384 H.M. HASSAN
oxide dismutase in glucose-limited chemostat cultures of Esch-
erichia coli. J. Bacteriol. 130:805-811; 1977.
26. Hassan, H. M.; Fridovich, I. Regulation of superoxide dis-
mutase synthesis in Escherichia coli: Glucose effect. J. Bac-
teriol. 132:505-510; 1977.
27. Hassan, H. M.; Fridovich. I. Regulation of the synthesis of
superoxide dismutase in Escherichia coli: induction by methyl
viologen. J. Biol. Chem. 252:7667-7672; 1977.
28. Hassan, H. M.; Fridovich. I. Superoxide radical and the ox-
ygen enhancement of the toxicity of paraquat in Escherichia
coli. J. Biol. Chem. 253:8143-8148; 1978.
29. Hassan, H. M.; Fridovich, I. Superoxide, hydrogen peroxide,
and oxygen tolerance of oxygen-sensitive mutants of Esche-
richia coli. Rev. Infect. Dis. 1:357-369; 1979.
30. Hassan, H. M.; Fridovich, I. Superoxide dismutase andits role
for survival in the presence of oxygen. In: Shilo, M., ed.
Strategy of microbial life in extreme environments. New York:
Verlag Chemie; 1979:179-193.
31. Hassan, H. M.; Fridovich, I. Paraquat and Escherichia coli:
mechanism of production of extracellular superoxide radical.
J. Biol. Chem. 254:10846-10852; 1979.
32. Hassan, H. M.; Fridovich, I. The mechanism of the antibiotic
action of pyocyanine. J. Bacteriol. 141:156-163; 1980.
33. Hassan, H. M.; Fridovich, I. Exacerbation of oxygen toxicity
by redox active compounds. In: King, T. E.; Mason, H. S.;
Morrison, M., eds. Oxidases and related redox systems. Bal-
timore, MD: University Park Press; 1982:151-167.
34. Hassan, H. M.; Moody, C. S. Superoxide dismutase protects
against the toxicity and mutagenicity of oxygen free radicals
in Salmonella typhimurium. Can. J. Physiol. Pharmacol. 60:82-
92; 1982.
35. Carlioz, A.; Touati, D. Isolation of superoxide dismutase mu-
tants in Escherichia coil: is superoxide dismutase necessary
for aerobic life? EMBO J. 5:623-630; 1986.
36. Touati, D. Molecular genetics of superoxide dismutases. Free
Radic. Biol. & Med. 5:393-402.
37. Natvig, D. O.; Imlay, K.; Touati, D.; Hallewell, R. Human
copper-zinc superoxide dismutase complements superoxide
dismutase deficient E. coli mutants. J. Biol. Chem. 262:14697-
14701; t987.
38. Elroy-Stein, O.; Bernstein, Y.; Groner, Y. Overproduction of
human Cu/Zn-superoxide dismutase in transfected cells: ex-
tenuation of paraquat-mediated cytotoxicity and enhancement
of lipid peroxidation. EMBO J. 5:615-622; 1986.
39. Krall, J.; Bagley, A. C.; Mullenbach, G. T.; Hallewell, R.
A.; Lynch, R. E. Superoxide mediates the toxicity of paraquat
for cultured mammalian cells. J. Biol. Chem. in press; 1988.
40. Eliasson, R.; Jornvall, H.; Reichard, P. Superoxide dismutase
participates in the enzymatic formation of the tyrosine radical
of ribonucleotide reductase from Escherichia coli. Proc. Natl.
Acad. Sci. USA 83:2373-2377; 1986.
41. Fontecave, M.; Griislund, A.; Reichard, P. The function of
superoxide dismutase during the enzymatic formation of the
free radical of ribonucleotide reductase. J. Biol. Chem.
262:12332-12336; 1987.
42. Kuo, C. F.; Mashino, T.; Fridovich, I. et,13-Dihydroxyisoval-
crate dehydrogenase: a superoxide-sensitive enzyme. J. BioL
Chem. 262:4724-4727; 1987.
43. Koao, Y.; Fridovich. I. Superoxide radical inhibits catalase.
J. Biol. Chem. 257:5751-5754; 1982.
44. Blum, J.; Fridovich, I. Inactivation of glutathione peroxidase
by superoxide radical. Arch. Biochem. Biophys. 240:500-508;
1985.
45. Hassan, H. M.; Fridovich, I. Superoxide dismutases: detoxi-
cation of a free radical. In: Jakoby, W. B., ed. Enzymatic basis
ofdetoxication. Vol. 1. New York: Academic Press; 1980:311-
332.
46. Hassan, H. M.; Fridovich, I. Enzymatic defenses against the
toxicity of oxygen and of streptonigrin in Escherichia coll. J.
Bacteriol. 129:1574-1583; 1977.
47. Dougherty, H. W.; Sandowski, S. J.; Baker, E. E. A new iron-
containing superoxide dismutase from Escherichia coli. J. Biol.
Chem. 253:5220-5223: 1978.
48. Clare, D. A.; Blum, J.; Fridovich, I. A hybrid superoxide
dismutase containing both functional iron and manganese. J.
Biol. Chem. 259:5932-5936; 1984.
49. Hassan, H. M.; Moody, C. S. Regulation of manganese-con-
taining superoxide dismutase in Escherichia coli: anaerobic
induction by nitrate. J. Biol. Chem. 262:17173-17177; 1987.
50. Hassan, H. M. Superoxide and superoxide dismutases in Esch-
erichia coli: threat and defense. In: Bloch, K.; Bolis, L.; Toste-
son, D. C., eds. Membranes, molecules, toxins and cells.
Boston: John Wright, PSG Inc.; 1981:123-133.
51. Gregory, E. M.; Fridovich. I. Induction of superoxide dis-
mutase by molecular oxygen. J. Bacteriol. 114:543-548; 1973.
52. Schiavone, J. R.; Hassan, H. M. Biosynthesis of superoxide
dismutase in eight prokaryotes: effects of oxygen, paraquat
and an iron chelator. FEMS Microbiol. Lett. 42:33-38; 1987.
53. DiGuiseppi, J.; Fridovich, I. Oxygen toxicity in Streptococcus
sanguis: the relative importance of superoxide and hydroxyl
radicals. J. Biol. Chem. 257:4046-4051; 1982.
54. Stowers, M. D.; Elkan, G. H. An inducible iron-containing
superoxide dismutase in Rhizobium japonicum. Can. J. Mi-
crobiol. 27:1202-1208; 1981.
55. Von Stein, R. S.; Barber, L. E.; Hassan, H. M. Biosynthesis
of oxygen-detoxifying enzymes in Bdellovibrio stolpii. J. Bac-
teriol. 152:792-796; 1982.
56. Hassan, H. M.; Fridovich, I. Intracellular production of su-
peroxide radical and of hydrogen peroxide by redox active
compounds. Arch. Biochem. 196:385-395; 1979.
57. Fridovich, I.; Hassan, H. M. Paraquat and the exacerbation
of oxygen toxicity. Trends in Biochem. Sci. 4:113-115; 1979.
58. Hassan, H. M.; Moody, C. S. Induction of the manganese-
containing superoxide dismutase in Escherichia coli by nali-
dixic acid and by iron chelators. FEMS Microbiol. Lett. 25:233-
236; 1984.
59~ Moody, C. S.; Hassan, H. M. Induction of manganese super-
oxide dismutase in Escherichia coli by iron chelators. Fed.
°Proc. 43:1759 abstr. (1984).
60. Moody, C. S.; Hassan, H. M. Anaerobic biosynthesis of the
manganese-containing superoxide dismutase in Escherichia coll.
J. Biol. Chem. 259:12821-12825; 1984.
61. Schiavone, J. R.; Hassan, H. M. The role of redox in the
regulation of manganese-containing superoxide dismutase bio-
synthesis in Escherichia coli. J. Biol. Chem. 263 in press:
1988.
62. Whiteside, C.; Hassan, H. M. Induction and inactivation of
catalase and superoxide dismutase of Escherichia coil by ozone.
Arch. Biochem. Biophys. 257:464-471; 1987.
63. laugh, S. Y.; Fridovich, I. Induction of superoxide dismutases
and hydroperoxidases in Escherichia coli B BI2 (ATCC #29682)
by manganese, iron and chelating agents. Fed. Proc. 43 3721
abstr. (1984).
64. Pugh, S. Y.; DiGuiseppi, J. L.; Fridovich, I. Induction of
superoxide dismutases in Escherichia coli by manganese and
iron. J. Bacteriol. 160:137-142; 1984.
65. laugh, S. Y.; Fridovich, I. Induction of superoxide dismutases
in Escherichia coli by metal chelators. J. Bacteriol. 162:196-
202; 1985.
66. Takeda, Y.; Avila, H. Structure and gene expression of the
Escherichia coli Mn-superoxide dismutase gene. Nucleic Acid
Res. 14:4577-4589; 1986.
67. Touati, D.; Carlioz, A. Superoxide dismutase mutants of E.
coli. In: Rotilio, G., ed. Superoxide and superoxide dismutase
in chemistry, biology and medicine. North Holland: Elsevier
Science Publisher; 1986:287-292.
68. Touati, D. Transcriptional and post-transcriptional regulation
of MnSOD biosynthesis in E. coli: a study using operon and
protein fusions. J. Bacteriol. in press; 1988.
69. Nettleton, C. J.; Bull, C.; Baldwin, T. O.; Fee, J. A. Isolation
of the Escherichia coli iron superoxide dismutase gene: evi-
dence that intracellular superoxide concentration does not reg-
 Control of SOD biosynthesis
ulate oxygen-dependent synthesis of the manganese superox-
ide dismutase. Proc. Natl. Acad. Sci. USA 81:4970-4973;
! 984.
70. Kirby, T.; Blum, J.; Kahane, I.; Fridovich, 1. Distinguishing
between Mn-containing and Fe-containing superoxide dismu-
tase in crude extracts of cells. Arch. Biochem. Biophys. 201:55 i -
555; 1980.
71. Ose, D. E.; Fridovich, I. Superoxide dismutase: reversible
removal of manganese and its substitution by cobalt, nickel,
or zinc. J. Biol. Chem. 2,51:1217-1218; 1976.
72. Ose, D. E.; Fridovich, I. Manganese-containing superoxide
dismutase from Escherichia coil: reversible resolution and metal
replacement. Arch. Biochem. Biophys. 194:360-364; 1979.
73. Gregory, E. M.; Dapper, C. H. Isolation of iron-containing
superoxide dismutase from Bacteroidesfragilis: reconstitution
as a Mn-containing enzyme. Arch. Biochem. Biophys. 220:293-
300; 1983.
74. Gregory, E. M. Characterization of the O2-induced man-
ganese-containing superoxide dismutase from Bacteroidesfra-
gills. Arch. Biochem. Biophys. 23&83-89; 1985.
75. Meier, B.; Barra, D.; Bossa, E; Calabrese, L.; Rotilio, G.
Synthesis of either Fe- or Mn-superoxide dismutase with an
apparently identical protein moiety by an anaerobic bacterium
dependent on the metal supplied. J. Biol. Chem. 257:13977-
13980; 1982.
76. Martin, M. E.; Byers, B. R.; Olson, M. O.J.; Salin, M. L.;
Arceneaux, J. E.L.; Tolbert, C. A Streptococcus mutans su-
peroxide dismutase that is active with either manganese or iron
as a cofactor. J. Biol. Chem. /,61:9361-9367; 1986.
77. Little, J. W.; Mount, D. W. The SOS regulatory system of
Escherichia coll. Cell 29:11-22; 1982.
78. Neidhardt, E C.; Varbogelen, R. A.; Vaughn, V. The genetics
and regulation of heat shock proteins. Ann. Rev. Genet. 18:295-
329; 1984.
79. Christman, M. F.; Morgan, R. W.; Jacobson, F. S.; Ames, B.
N. Positive control of a regulon for defenses against oxidative
stress and some heat shock proteins in Salmonella typhimu-
rium. Cell 41:753-762; 1985.
80. Brawn, K.; Fridovich, I. Strand scission by enzymically gen-
erated oxygen radicals. Arch. Biochem. Biophys. 206:414-
419; 1981.
81. Brawn, M. K.; Fridovich, I. Increased superoxide radical pro-
duction evokes inducible DNA repair in Escherichia coll. J.
Biol. Chem. 260:922-925; 1985.
82. Chan, E.; Weiss, B. Endonuclease IV of Escherichia coil is
induced by paraquat. Proc. Natl. Acad. Sci. USA 84:3189-
3193; 1987.
83. Farr, S. B.; Natvig, D. O.; Kogoma, T. Toxicity and muta-
genicity of plumbagin and the induction of a possible new DNA
repair pathway in Escherichia coll. J. Bacteriol. 164:1309-
1316; 1985.
84. Hancock, L.; Hassan, H. M. Regulation of the manganese-
containing superoxide dismutase is independent of the indu-
cible DNA repair system in Escherichia coll. J. Biol. Chem.
260:12954.-12956; 1985.
85. Privalle, C. T.; Fridovich, I. Induction of superoxide dismutase
in Escherichia coli by heat shock. Proc. Natl. Acad. Sci. USA
84:2723-2726; 1987.
86. Kogoma, T.; Farr, S. B.; Joyce, K.; Natvig, D. O. Isolation
View publication stats
385
of operon fusions (soi-lac Z) inducible by oxidative stress in
E. coll. ln: UCLA symposium mechanisms and consequences
of DNA damage processings. Abstr; 1988.
87. Touati, D. Cloning and mapping of the manganese superoxide
dismutase gene (sad A) of Escherichia coil K-12. J. Bacteriol.
155:1078-1087; 1983.
88. Sakamoto, H.; Touati, D. Cloning of the iron superoxide dis-
mutase gene (sod B) in Escherichia coil K-12. J. Bacteriol.
159:418-420; 1984.
89. Carlioz, A.; Ludwig, M. L.; Stallings, W. C.; Fee, J. A.;
Steinman, H. M.; Touati, D. Iron superoxide dismutases: nu-
cleotide sequence of the gene from E. coli K- 12 and correlation
with crystal structures. J. Biol. Chem. in press; 1988.
90. Lee, F. J.; Hassan, H. M. Biosynthesis of superoxide dismutase
and catalase in Saccharomyces cerevisiae: effects of oxygen
and cytochrome-e deficiency. J. Industr. Microbiol. 1:187-
193; 1986.
91. Lee, F. J.; Hassan, H. M. Biosynthesis of superoxide dismutase
in Saccharomyces cerevisiae: effects of paraquat and copper.
J. Free Radicals Biol. Med. 1:319-325; 1986.
92. Lee, F. J.; Hassan, H. M. Biosynthesis of superoxide dismutase
and catalase in chemostat culture of Saccharomyces cerevisiae.
Appl. Microbiol. Biotech. 26:531-536; 1987.
93. Bettger, W. J.; Savage, J. E.; O'Dell, B. L. Effects of dietary
copper and zinc on erythrocyte superoxide dismutase activity
in the chick. Nutr. Rep. int. 19:893-900; 1979.
94. Ljutakova, S. G.; Russanov, E. M.; Liochev, S. I. Copper
increases superoxide dismutase activity in rat liver. Arch.
Biochem. Biophys. 235:636-643; 1984.
95. Statzman, A. R.; Kosman, D. J. The utilization of copper and
its role in the biosynthesis of copper-containing proteins in the
fungus, Dactylium dendroides. Biochim. Biophys. Acta 544:163-
179; 1978.
96. Gregory, E. M.; Goscin, S. A.; Fridovich, I. Superoxide and
oxygen toxicity in a eukaryote. J. Bacteriol. 117:456-460;
1974.
97. Crape, J. D.; Tierney, D. L. Superoxide dismutase and pul-
monary oxygen toxicity. Am. J. Physiol. 2,2,6:1401-1407; 1974.
98. Stevens, J. B.; Autor, A. P. Induction of superoxide dismutase
in neonatal rat lung. J. Biol. Chem. 2,52:3509-3514; 1977.
99. Housset, B.; Junod, A. E Enzyme response of cultured en-
dothelial cells to hyperoxia. Bull. Eur. Physiopathol. Respir.
17(suppl.):107-110; 1981.
100. Scandalios, J. G. The antioxidant enzyme genes Cat and Sod
of maize: regulation, functional significance, and molecular
biology. In: Rattazzi, M. C.; Scandalais, J. G.; Whitt, G. S.,
eds. isozymes: current topics in biological and medical re-
search. Vol. 14. New York: Alan R. Liss; 1987:19-44.
101. Tanaka, K.; Sugahara, K. Role of superoxide dismutase in
defense against sulfur dioxide toxicity and an increase in su-
peroxide dismutase activity with sulfur dioxide fumigation.
Plant Cell Physiol. 21:601-612; 1980.
102. Matters, G. L.; Scandalios, J. G. Effect of the free radical-
generating herbicide paraquat on the expression of the super-
oxide dismutase (Sod) genes in maize. Biochim. Biophys. Acta
882:29-38; 1986.
103. Matters, G. L.; Scandalios, J. G. Effect of elevated temper-
ature on catalase and superoxide dismutase during maize de-
velopment. Differentiation 30:190-196; 1986.