Transcript The Cell, 5e

Chapt. 24 Oxygen toxicity
Ch. 24 Oxygen toxicity,
free-radical injury
Student Learning Outcomes:
• Explain how O2 is both essential to life and toxic
•
Oxidative phosphorylation, Reactive Oxygen radicals
• Explain formation of the major ROS
•
Enzymatic by products, non-enzymatic
• Describe toxic effects of ROS on cells
• Describe nitrogen NO and RNOS radicals
• Explain cell protective mechanisms
•
Enzymes, antioxidants, compartmentalization
• Describe association between ROS and diseases
Table 1 diseases associated with free-radical injury
Table 1 diseases associated with free-radical injury:
• Amyotrophoic lateral sclerosis (ALS)
• Ischemia/reperfusion injury
• OXPHOS diseases (mitochondria)
• Alzheimer’s disease
• Parkinson’s disease
• Diabetes
• Aging
Fig. 2; Cell has defenses against
damage by ROS and RNOS
(Reactive oxygen species,
Reactive nitrogen-oxygen species):
Antioxidants, enzymes
Reactive Oxygen Species
Reactive Oxygen Species (ROS) are a natural
occurrence:
• Accidental products of nonenzymatic
and enzymatic processes
• Deliberate production by immune
cells killing pathogens
• UV irradiation, pollutants
• Cells have many defenses
Fig. 1 O2 has 2 antibonding
e- with parallel spins;
tendency to form toxic ROS
Radical nature of Oxygen
Radical nature of oxygen:
• Radical is an unpaired e• Free radical has independent
existence (not bound to enzyme)
• Free radical extracts e- from other
molecules
• O2- accepts e- from strong reducer
such as CoQH• in ETC
Fig. 3, O2 can accept
total of 4 e- to form H2O
Table 2 ROS species
Table 2 Some Reactive Oxygen species (ROS)
O2-
Superoxide anion Produced by ETC and other sites;
does not diffuse far, generates other ROS, such
as by reaction with H2O2 in Haber-Weiss reaction
H2O2 Hydrogen peroxide Not a free radical, but generates free
radicals by reaction with transition metal (e.g. Fe2+);
diffuses into and through cell membranes
OH•
Hydroxyl radical
The most reactive in attacking
biological molecules; produced from H2O2
in Fenton reaction in presence of Fe2+ or Cu+
HOCl Hypochlorous acid Produced from H2O2 by neutrophils
to destroy invading organisms (OCl-)
Hydroxyl radical
Formation of very reactive oxygen species OH•
• Two nonenzymatic reactions can form OH• by
transfer of single e• Metals Fe2+ or Cu+ are kept sequestered
Fig. 4
C. Major sources of ROS
C. Major source of ROS:
• COQH• in electron transport chain
accidental interaction with O2
(COQH• is free in membrane)
Fig. 5; ETC
Fe-H is Fe-heme of cytochromes
Sources of ROS
Oxidases, oxygenases, peroxidases generate ROS:
• Enzymes bind O2, transfer 1 e- via metals;
• Accidental leakage of free-radical intermediates
• Ex. Cyt P450 mono-oxygenases detoxify many organic
compounds (alcohol, drugs, toxic chemicals like CCl4)
• Peroxidases generate H2O2 (ex. VLCFA in peroxisomes)
Fig. 19.12, 19.13
cytochrome P450
mono-oxygenase
Sources of ROS
Ionizing Radiation generates OH•
• Cosmic rays
• Radioactive chemicals
• X-rays
May also generate organic radicals
from contact biomolecules
Fig. 6
Oxygen radicals react with cell components
Oxygen radicals react with cell components:
•
•
•
•
Lipid peroxidation of membranes
Increased permeability → influx Ca2+ → mitochondrial damage
Cys SH and other aa of proteins oxidized and degraded
DNA oxidized → breakage
Fig. 7*
Lipid peroxy radicals
Lipid peroxidation: free-radical chain reaction:
A. Initiation by OH• attack of poylunsaturated lipid → lipid•
B. free-radical chain reaction by reaction with O2
C. Lipid peroxy radical propagates, lipid peroxide degrades
D. Terminate by vitE or lipid-soluble antioxidants
Major contribution to
ROS-induced injury
Fig. 8
ROS attack proteins, peptides, DNA
ROS attack proteins, peptides and DNA
• Pro, his, arg, cys, met most susceptible
• Protein fragment, cross-link, may aggregate, also
will be degraded
• Glutathionine (g-glu-cys-gly) is
anti-oxidant, cell defense
• DNA oxidized bases mispair
at replication (G-C → T-A)
• DNA backbone broken
• repair mechanisms exist
Fig. 9
Reactive Nitrogen-oxygen species
Nitric Oxide and Reactive Nitrogen-oxygen species
NO is both essential for life and toxic:
• Gas, diffuses through membranes
• Low concentrations: neurotransmitter, hormone (vasodilation)
• Nitroglycerin tablets release NO, vasodilator for heart
• Binds Fe-heme in receptor guanylyl cyclase,
cGMP activates signal cascade
NO works short distance from source
• nNOS and eNOS regulated by Ca2+
• iNOS inducible in immune cells,
prouces high levels of NO
Fig. 10
NO is toxic
At high concentrations,
NO is toxic, RNOS form
• RNOS can cause as much
damage as ROS, plus also
do nitrating, nitrosylating
• RNOS damage proteins,
cause lipid peroxidation,
DNA breaks
RNOS are involved in:
• neuro-degenerative
diseases like Parkinson’s,
• chronic inflammation like
rheumatoid arthritis (RA)
Fig. 11
Phagocytosis uses free radicals
Phagocytic cells of immune system do respiratory
burst: O2 → ROS, RNOS
•
•
•
•
Part of antimicrobial defense, also anti-tumor (~ 30-60 min)
NADPH oxidase forms O2- → H2O2 and OH•
Myeloperoxidase forms HOCl → OCliNOS activated, makes NO → RNOS
Free-radical release in
some disease states
contributes to injury;
chronic inflammation
Fig. 12
V. Cellular Defenses
V. Cells have defenses against oxygen toxicity:
• Antioxidant scavenging enzymes (red)
• Nonenzymatic antioxidants (free radical scavengers)
• Compartmentalization
• Repair of damaged components
• Metal sequestration
Fig. 13* compartmentalization
SOD = superoxide dismutase
GSH = glutathione
Antioxidant scavenging enzymes
Antioxidant scavenging enzymes:
• Superoxide dismutase (SOD)
• Converts O2- to H2O2
• 3 isoforms:

Cytosol, mitochondria, extracellular
• Catalase
• Reduces H2O2 to H2O
• Prevents OH• formation
• mostly peroxisome
Fig. 14
Antioxidant enzymes
• Glutathione peroxidase, glutathione reductase:
•
GSH = glutathione (g-glu-cys-gly)
• Peroxidase reduces H2O2, oxidizes two SH groups → GSSG
• Peroxidase in cytosol, mitochondria, have selenium
• Reductase recycles the glutathione, reduces with NADPH
Fig. 15
Nonenzymatic antioxidants
Vitamin E (a-tocopherol) is antioxidant:
• Lipid-soluble, protects against lipid peroxidation in
membranes
• Nonenzymatic terminator of free-radical chain reaction
• Lipid fraction of vegetable oils,
liver, egg yolks, cereals
• Lipoprotein particles in blood
Fig. 16
Antioxidants
Vitamin C (ascorbate) is antioxidant:
• Can donate e- to vitamin E to regenerate Vitamin E
• Water-soluble, circulates blood and fluids to access
membranes
• Vitamin C is also redox coenzyme for collagen synthesis,
other reactions
Fig. 17
Antioxidants
Carotenoids (b-carotene, precursor of vitamin A):
• Are antioxidants, found in fruits and vegetables
• May slow cancer, atherosclerosis
• Lutein and zeoxanthin in macula of eye (may help protect
against macular degeneration of retina)
Fig. 18
Antioxidants
Flavonoids are antioxidants:
• May inhibit enzymes responsible for ROS (xanthing oxidase)
• May chelate Fe and Cu; free-radical scavengers
Endogenous antioxidants: melatonin and uric acid
Fig. 19,20
Parkinson’s disease and neuronal degeneration
Model for ROS and RNOS in
neuronal degradation in
Parkinson’s disease:
• Dopamine levels are reduced
because degenerated dopaminergic
neurons
• Unknown trigger, model:
• MAO (monoamine oxidase)
generates H2O2 → (also SOD in Mt)
• Damaged Mt leak Fe2+ → OH•
• NO forms RNOS
• Radical chain reactions
Fig. 21
Protection against ozone in lung epithelium
Protection against ozone in lung lining fluid:
• Many pathways to protect from ozone, which can form
ROS
Fig. 22
AA = vit C
GSH-Px = perosidase
Neutrophil activation
can worsen problem
Key Concepts
Key concepts:
• Oxygen radical generation contributes to cell death and
degeneration in a variety of diseases
• Radical damage occurs via e- extraction from biologic
molecules
• ROS include superoxide, hydrogen peroxide, hydroxyl radical
• ROS can damage DNA, proteins, lipids, lead to cell death
• Other radical species are NO and HOCl
• NO reacts with oxygen or superoxide to form RNOS
• The immune response produces radical species to destroy
microorganisms (superoxide, HOCl, NO)
• Cell defenses against radical damage include defense
enzymes, antioxidants and compartmentalization.
Review question
5. The level of oxidative damage to mitochondrial DNA is 10
times greater than that to nuclear DNA. This could be due, in
part, to which of the following?
A. Superoxide dismutase is present in the mitochondria
B. The nucleus lacks glutathione
C. The nuclear membrane presents a barrier to ROS
D. The mitochondrial membrane is permeable to ROS
E. Mitochondrial DNA lacks histones