Antibodies Produce Ozone During Bacterial Killing and Inflammation
By Jason Socrates
Bardi
The Scripps Research Institute (TSRI) President Richard
A. Lerner, Associate Professor Paul Wentworth, Jr., Ph.D.,
and a team of investigators at TSRI is reporting that antibodies
can destroy bacteria, playing a hitherto unknown role in immune
protection. Furthermore, the team found that when antibodies
do this, they appear to produce the reactive gas ozone.
"[Ozone] has never been considered a part of biology before,"
says Lerner, who is Lita Annenberg Hazen Professor of Immunochemistry
and holds the Cecil H. and Ida M. Green Chair in Chemistry.
The report will appear in an upcoming issue of the journal
Science.
The ozone may be part of a previously unrecognized killing
mechanism that would enhance the defensive role of antibodies
by allowing them to subject pathogens to hydrogen peroxide
and participate directly in their killing. Previously, antibodies
were believed only to signal an immune response.
This ability of antibodies to generate toxic compounds may
also link them to a number of inflammatory diseases, such
as atherosclerosis, lupus, multiple sclerosis, and rheumatoid
arthritis. Furthermore, this research opens up exciting possibilities
for new antibody-mediated therapies for conditions ranging
from bacterial and viral infection to cancer.
Recognition and Killing in the Same Molecule
Also called immunoglobulins, antibodies are secreted proteins
produced by immune cells that are designed to recognize a
wide range of foreign pathogens. After a bacterium, virus,
or other pathogen enters the bloodstream, antibodies target
antigens—proteins, fat molecules, and other pieces of
the pathogen—specific to that foreign invader. These
antibodies then alert the immune system to the presence of
the invaders and attract lethal "effector" immune cells to
the site of infection.
For the last hundred years, immunologists have firmly held
that the role of antibodies was solely to recognize pathogens
and signal the immune system to make an immune response. The
conventional wisdom was that the dirty work of killing the
pathogens was to be left to other parts of the immune system.
Now, Lerner, Wentworth, and their colleagues have demonstrated
that antibodies also have the ability to kill bacteria. This
suggests that rather than simply recognizing foreign antigens
and then activating other parts of the immune system to the
site of infection, the antibodies may further enhance the
immune response by directly killing some of the bacteria themselves.
Antibodies do this by producing the chemical oxidant hydrogen
peroxide—best known as the foamy formulation used for
first-aid. Hydrogen peroxide is lethal to bacterial cells
because it pokes holes in their cell walls, bursting the cells
and killing them.
In the Science paper, the TSRI team reports the effective
killing of E. coli bacteria through hydrogen peroxide production
by antibodies specific for that bacteria.
The Ozone Hole in Each One of Us
Certainly the most surprising result that Lerner, Wentworth,
and their colleagues found was that antibodies appear to make
ozone, which they detected through its chemical signature.
They have not yet demonstrated conclusively that what they
found is ozone, but they are highly confident that ozone is
what the antibodies are producing because no other known molecule
has the same chemical signature.
Ozone is a particularly reactive form of oxygen that exists
naturally as a trace gas in the atmosphere, constituting on
the average fewer than one part per million air molecules.
But it is noted mainly where its presence or absence poses
a threat to public health.
The gas is perhaps better known for its crucial role absorbing
ultraviolet radiation in the upper reaches of earth's stratosphere—about
25 km above the surface—where it is concentrated in a
so-called ozone layer, protecting life on earth from damaging
solar radiation.
Ozone is also a familiar component of air in industrial
and urban settings where the highly reactive gas is a hazardous
component of smog in the summer months. Never before has ozone
been detected in biology.
"All our analytical data point to this oxidant possessing
the chemical signature of ozone," says Wentworth, "in which
case, this is a new molecule in biology and therefore may
have tremendous ramifications for signaling and inflammation."
Proof for a Proposed Reaction Pathway
All antibodies have the ability to produce hydrogen peroxide,
the report adds, but they need to first have available a molecule
known as "singlet" oxygen—another highly reactive oxygen
species—to use as a substrate.
Singlet oxygen is an electronically excited form of oxygen
that forms spontaneously during normal metabolic processes
or when oxygen is subjected to visible or ultraviolet light
in the presence of a sensitizer. "Phagocytic" innate immune
cells, like neutrophils, also produce singlet oxygen and are
the most likely source of the substrate for antibodies, since
during an immune response, antibodies will recruit neutrophils
and other immune cells to the site of an infection.
Once there, the neutrophils will engulf and destroy bacteria
and other pathogens by blasting them with singlet oxygen and
other oxidative molecules. The antibodies reduce singlet oxygen
by combining it with water to produce hydrogen peroxide, producing
ozone as a side product.
Interestingly, all antibodies have the ability to do this,
which leads the TSRI team to speculate that the removal of
singlet oxygen may have been the original role of antibodies.
In a previous report, the same team speculated an ancient
form of antibodies may have existed—molecules whose role
was to catalyze singlet oxygen destruction, since singlet
oxygen can potentially destroy any cell, making it dangerous
to have around. Prior to the evolution of the modern antibody-mediated
humoral immune response in vertebrates hundreds of millions
of years ago, ancient antibodies may have been responsible
for controlling the release of highly reactive and potentially
dangerous singlet oxygen. Later, when antibodies developed
as part of the adaptive arm of the immune system, they kept
their original function because it provided a bit of extra
lethality.
Another interesting finding is that the antibodies carry
the reaction through an unusual intermediate. Lerner, Wentworth,
and their colleagues postulate that the antibodies carry the
reaction through an intermediate chemical species of dihydrogen
trioxide, a reduced form of ozone.
Dihydrogen trioxide has also never before been observed
in biological systems, and its presence as an intermediate
has been the source of considerable speculation in the scientific
community.
The team's reported detection of ozone is strong support
of this proposed dihydrogen trioxide intermediate, and now
the team is tackling the larger question of what it means.
"This is a novel set of observations and very interesting
ones—there are a million questions [we could ask]," says
TSRI Professor Bernard Babior, "What does the ozone do to
the body's proteins, nucleic acids? Are there lethal concentrations
of ozone? Does it have anything to do with other reactive
species in the body?"
The research article, "Evidence for Antibody-Catalyzed Ozone
Formation in Bacterial Killing and Inflammation" is authored
by Paul Wentworth, Jr., Jonathan E. McDunn, Anita D. Wentworth,
Cindy Takeuchi, Jorge Nieva, Teresa Jones, Cristina Bautista,
Julie M. Ruedi, Abel Gutierrez, Kim D. Janda, Bernard M. Babior,
Albert Eschenmoser, and Richard A. Lerner, and appears in
the November 18, 2002 "Science Express," the advanced publication
edition of the journal Science. The article will appear
in Science later this year.
The research was funded by the National Institutes of Health,
The Skaggs Institute for Chemical Biology, and an A.R.C.S.
fellowship.
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