Reactive Oxygen-intermediates in biology and medicine- Lecture 72

Question 1

What is a free radical?

Answer 1

a species which contains an unpaired electron in its outer orbital
very reactive

Question 2

What diseases have ROS been implicated in?

Answer 2

neurodegenerative diseases (CNS- MAO, high levels of Fe)
cancer (DNA damage, mutations)
free radical theory of aging
foam cells and cardiovascular disease
smoking and lung injury
reperfusion and transplant injury
Others (sepsis, inflammation, oxygen toxicity, radiation injury, etc.)

Question 3

What is oxidative stress?

Answer 3

imbalance between the prodoxidant and antioxidant levels in the cells and tissues

Question 4

_____ are ROS which can cause tissue damage and whose levels may be increased by _____.

Answer 4

Prooxidants

certain drugs, infection, external exposures, tissue injury

Question 5

What is the function of antioxidants?

Answer 5

inhibit formation of FOS or remove/ scavenge the generated ROS

Question 6

What can cause oxidative stress?

Answer 6

increase in prooxidant formation
and/or
decrease/deficiency in antioxidant levels

Question 7

What is the triplet state of oxygen?

Answer 7

molecular oxygen in its ground state

2 unpaired electrons in its outer orbital (diradical) that are spinning in the same direction

Question 8

What is the singlet ground state of oxygen?

Answer 8

absorption of energy by triplet oxygen inverts the spin of one of the electrons to produce this
very reactive

Question 9

What can the two electrons of the reactive singlet ground state of oxygen insert into?

Answer 9

the double bonds of:
aromatic amino acids of proteins (Phe, Tyr, Try)
unsaturated fatty acids
the purine and pyrimidine bases of DNA and RNA
*this disrupts the normal structures and interactions

Question 10

____ is the most important reactive oxygen species in the biological system.

Answer 10

Superoxide anion radical

Question 11

Describe the dismutation reaction of superoxide.

Answer 11

superoxide + superoxide + 2H+ –> H2O2 + O2

via superoxide dismutase (SODs) OR can occur non-enzymatically

Question 12

What kinds of reactions involve free radicals?

Answer 12

Hydrogen abstraction
addition to double bond
termination
dismutaion/disproportionation
Haber-Weis Rxn
Fenton Rxn

Question 13

The _____ radical is the most powerful ROS produced in biological and chemical systems and can react with any biochemical or macromolecule.

Answer 13

OH

Question 14

Why is OH radical central to the toxicity produced by ROS.

Answer 14

it reacts and inactivates or disrupts proteins, lipids, DNA, and RNA

Question 15

Describe the Haber-Weis reaction.

Answer 15

H2O2 + superoxide –> O2 + OH- + OH(radical)

slow thermodynamically if not catalyzed by iron or copper

Question 16

Describe the Fenton reaction.

Answer 16

H2O2 + Fe2+ –> OH(radical) + OH- + Fe3+

Fe3+ + superoxide –> Fe2+ + O2

Question 17

Name the systems that produce ROS.

Answer 17

mitochondrial respiratory chain (complexes I and III)
NADPH oxidases
cytochrome P450s-Cyp2E1
reprofusion injury (xanthine dehydrogenase --> xanthine oxidase)
oxidative enzymes
autooxidizable biochemicals
autooxidizable hemes
external agents
metal overload

Question 18

What kinds of oxidative enzymes produce ROS?

Answer 18

monoamide oxidase, urate, glycolate, D-aa, oxidases, peroxisomal fatty acyl CoA, prostaglandin synthesis

Question 19

Why are ROS toxic?

Answer 19

interactions with proteins
interaction with lipids
interaction with DNA

Question 20

Why does ROS interactions with proteins result in toxicity ?

Answer 20

oxidized SH groups
oxidized methionine to methionine sulfoxide
fragment aromatic rings (Phe and Tyr) or indole ring of Try to carbonyls
nitration of tyrosine residues
disrupt peptide bonds
disrupt covalent bonds and non-covalent interactions

Question 21

Why does ROS interaction with lipids result in toxicity?

Answer 21

form lipid radicals and lipid hydroperoxides
destroy cellular membranes
form reactive aldehydes
form protein-aldehyde adducts

Question 22

Why do ROS interaction with RNA/DNA result in toxicity?

Answer 22

cleave phosphodiester bonds
cause base excision
oxidize bases
all result in base mispairing

Question 23

Describe the lipid peroxidation reaction.

Answer 23

Initiation:
LH --> L(radical)
Propagation:
L(radical) + O2 --> LOO(radical)
LOO(radical) + LH --> LOOH + L(radical)
Degradation:
LOOH --> malondialdehyde (MDA) + 4-hydroxynonenal (4-HNE)

Question 24

How can you halt the propagation step of the lipid peroxidation reaction?

Answer 24

Vitamin E (alpha-tocopherol) is a major antioxidant that reacts with lipid and lipid alkoxyl radical s to restore the lipid and form a vitamin E radical
can be regenerated by ascorbic acid (Vitamin C)

Question 25

What are the beneficial effects of ROS and free radicals?

Answer 25

activation of certain txn factors (eg. activation of Nrf2, NF-kB, AP1 via H2O2)
macrophages and neutrophils release ROS as a weapon against immunologic targets
enzymatic reactions

Question 26

How do phagocytes use ROS?

Answer 26

NADPH oxidase (produces superoxide) reactions 
myeloperoxide reactions (produce bleach)

Question 27

Describe chronic granulomatous disease.

Answer 27

failure to promote a respiratory burst largely due to deficiencies in some of the subunits which make up the NADH oxidase

Question 28

What mechanisms are in place to protect the body from damage from ROS.

Answer 28

superoxide dismutase
catalase
peroxidases
GSH + Gluthathione peroxidase (SE) + Gluthathione reductase (G6PDH generates NADPH for reduction of GSSG)
Glutathione transferase
metallothionein, ferritin ceruloplasmin
alpha-tocopherol (vitamin E), ascorbate (vitamin C), uric acid, caretenoids, GSH, SAM
heme oxygenase

Question 29

Describe the process of reperfusiontransplant injury.

Answer 29

when ATP is no longer being synthesized (due to ischemia of the tissue), it is converted to hypoxanthine which can be converted to xanthine via xanthine oxidase (which is converted from xanthine dehydrogenase when large amounts of hypoxanthine accumulates)
when O2 is re-added during reprofusion, you get injury of the tissue because it can further be converted to ROS

Question 30

How can reperfusion/transplant injuries be prevented?

Answer 30

addition of buffers containing protectants (antioxidants) + chelators + a xanthine oxidase inhibitor (normally allopurinol) with the reperfusion of O2

Question 31

Why does our mitochondria have the most ROS?

Answer 31

auto-oxidation of quinones
CoQ + E- + H+ --> semiquinone radical 
semiquinone radical + e- + H+ --> ubiquinol (CoQH2)
OR 
semiqinone + O2 --> CoQ + O2(radical)