Transcript Metals-and-Oxidative-Stress

Metals and Oxidative/Nitrosative Stress
oxidative/nitrosative stress: high levels of ROS (Reactive Oxygen
Species) and RNS (Reactive Nitrogen Species) such as free radicals
and peroxides which damage cellular components (DNA, proteins,
lipids)
NO,
O2 - , OH, ONOO(H), H2O2 , ROO, ROOH, HOCl, NO2-
Elevated ROS/RNS is linked to diseases associated with chronic
inflammation and aging e.g. atherosclerosis, neurological diseases,
cancer (Free Radical Theory of Aging)
A biochemical imbalance occurs between processes which produce
ROS/RNS and processes that destroy ROS/RNS
ROS/RNS: are essential in biological systems
e.g. cell signaling, immune response against pathogens
Antioxidants (enzymes and small molecules) play a key role in
controlling ROS/RNS levels
Nitric Oxide, NO (NO)
Roles in Biology include
 Produced in the immune response against pathogens
 Inducing relaxation in the smooth muscles lining the blood
vessels  vasodilation (binds to guanylate cyclase, a heme
enzyme)
 Inhibiting cell proliferation
 A neurotransmitter
 NO prodrugs used to treat
hypertension (high blood pressure)
angina (insufficient oxygen to the heart muscles)
nitroglycerin
Superoxide (O2•-)
 A byproduct of mitochondrial respiration and enzyme reactions
e.g. xanthine oxidase
 Immune system: the enzyme NADPH oxidase produces high
levels of O2•- to kill pathogens
 O2•- levels augmented in acute and chronic inflammation
Nitric oxide synthase can produce peroxynitrite under stress
conditions:
O2•- is a precursor of OH
Peroxynitrite, ONOO(H)
A strong oxidizing Eº (ONOO-, 2H+/NO2, H2O) = 1.6 V, pH 7,
nitrating and hydroxylation agent (especially for Tyr)
ONOOH spontaneously decomposes: (% yield at pH 7.4)
Synthesis of ONOO-:
Hydroxy Radical, •OH
One of the most potent ROS in biology since it reacts with all
biomolecules at essentially diffusion controlled rates ( 109 M-1 s-1)
OH
is generated from:
H2O2 (Fenton reaction)
peroxynitrite decomposition
O2 and H2O2 (Haber-Weiss reaction)
H2O ionization
Antioxidants: react rapidly (scavenge) ROS/RNS,
preventing their reactions with biomolecules
A) Important small molecule antioxidants: vitamin C, vitamin E,
carotenoids, thiols (glutathione, thioredoxin and lipoic acid),
flavonoids, melatonin
Example: glutathione scavenging of radicals (R•)
GSH + R• → GS• + RH
GS• + GS• → GSSG
– the ratio of GSH/GSSG is a good measure of oxidative
stress of an organisms
B) Antioxidant enzymes:
a) Superoxide dismutase (SOD)
b) Catalase
c) Glutathione peroxidase
a) Superoxide Dismutase, SOD
cytosolic Zn,Cu-SOD: k = 2 x 109 M-1 s-1
b) Catalase
found within the peroxisome; a heme protein
- one of the highest turnover rates for all enzymes: ∼6 x106 H2O2 /min
Metals Can Protect and Cause Oxidative/Nitrosative Stress
1. Free Metal Cations generate ROS in Biological
Systems via Redox Chemistry
Elevated levels of Fe2+/Fe3+ and/or Cu+, Cu2+ implicated in
many diseases associated with oxidative/nitrosative stress
hemochromatosis
Wilson’s disease
2. Metal Complexes can Prevent Oxidative/Nitrosative
Stress e.g. porphyrin complexes, cobalamins
Cobalamins as ROS/RNS Scavengers
There is considerable evidence that Cbl scavenges NO to form
NOCbl in biological systems
Evidence supporting NOCbl formation in vivo includes:
 NO inhibits the two mammalian B12-dependent enzymes (in
vitro and in cell studies)
 Cbl suppresses NO-induced relaxation of smooth muscle in
rodents, NO-induced vasodilation and NO-mediated inhibition
of cell proliferation
 Cbl reverses NO-induced neural tube defects
 Cbl regulates pro-inflammatory cytokines and growth factors
assoc. with the immune response
 Patients with severe Cbl deficiency have high TNF- levels and
low EGF levels which are corrected by Cbl replacement
 The transcobalamin receptor is upregulated by TNF-
 Inflammatory diseases associated with increased levels of
transcobalamin and its receptor
 Cbl depletion is associated with reversible immunodeficiency
and can promote HIV infection
 Cbl supplementation beneficial for wide variety of diseases
associated with oxidative stress and chronic inflammation
(chronic fatigue syndrome, trauma, sepsis, asthma, arthritis,
AD, MS, eczema)
Note: Cbl supplementation results in considerable amounts of free
(non-protein bound) Cbl in cells
Chemistry model studies show that the reaction between Cbl(II)
and NO is rapid:
Furthermore, expts have shown that Cbl(III)’s are reduced to Cbl(II)
in cells making the reaction catalytic
Can other ROS/RNS react with Cbl(II)?
Our work:
O2•- scavenger
k (M-1 s-1)
TEMPOL
3.4 x 105
FeIII tris[N-(2-pyridylmethyl)-2-aminoethyl]amine
2.2 x106
FeII tetrakis-N,N,N’,N’(2-pyridylmethyl)ethylendiamine
~ 3 x 107
MnIII meso-tetrakis(ortho-N-ethylpyridinium-2’-yl)
porphyrin
5.8 x 107
MnIII 5,10,15,20-tetrakis[N-methyl-N'-(2-methoxyethyl)
imidazolium-2-yl]porphyrin
9.5 x 107
MnIII 5,10,15,20-tetrakis[N-(2-methoxyethyl) pyridinium-2yl]porphyrin
1.1 x 108
MnIII tetrakis(N-(1-(2-(2-(2-methoxyethoxy)ethoxy)
ethyl)pyridinium -2-yl)porphyrin
1.3 x108
MnIII tetrakis(N,N'-di(1-(2-(2-(2-methoxyethoxy)
ethoxy)ethyl)imidazolium-2-yl)porphyrin
3.5 x 108
cob(II)alamin
5 x 108
M40401 (MnII porphyrin)
1.6 x 109
Cu, Zn-SOD
2 x 109