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Week 5 Flashcards

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1
Q

What is oxidative stress

A

Results form imbalance between the levels of reactive oxygen species and antioxidants

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2
Q

How can oxidative stress occur

A

If there is an excess level of ROS or a depleteion in AOX

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3
Q

What is an antioxidant

A

A molecule which protects a biological target against oxidative damage

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4
Q

What are the two groups of antioxidant

A

Enzymes (proteins) and low molecular weight non-protein organic molecules

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5
Q

What are the antioxidant enzymes

A

Catalases
Glutathione peroxidases
Peroxidases
Peroxiredoxins
SUperoxide dimutases
Thioredoxin

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6
Q

What are oxidants

A

Recative molecules that are produced both inside your body and the environment that can react with other cellular molecules such as protiens

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7
Q

What is the most common type of oxidant in the cells

A

Reactive Oxygen Species (ROS) it include Non-radicals and radicals

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8
Q

What are free radicals

A

ANy molecules containing one or more unpaired electrons

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9
Q

What are examples of non radicals

A

Hydrogen peroxide
Hypochlorous acid
Peroxynitrite
Singlet oxygen
ozone

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10
Q

What are some examples of radicals

A

Hydroxyl
Superoxide
Lipid peroxyl
Nitric oxide

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11
Q

What is oxygen in its ground state

A

Has two unpaired electrons with parallel spins (biradical)

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12
Q

What does oxygen being bi radical do

A

Unlikely to particpate in recations with stable organic molecules that are arranged in pairs with antiparallel spins unless oxygen is activated

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13
Q

How can oxygen be activated

A

If electrons are transferred to O2 one at a time producing mono-radical or non-radical molecules

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14
Q

What are the three types of ROS

A

Superoxide anion radical
Hydrogen peroxide
Hydroxyl radical

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15
Q

How are oxygen radicals formed

A

One electron reduction leads to the formation of superoxide anion radical then reductions of superoxide (addition of two electrons) produces hydrogen peroxide if it takes third electron you have hydroxyl radical

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16
Q

What happens once the hydroxyl radical is formed

A

can be reduced with an electron to from water (4 electron reduction of oxygen)

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17
Q

What are other ROS that can also be converted

A

Peroxynittrite (reaction of superoxide and nitric oxidie) and hypochlorous acid (hydrogen peroxide and chlorine)

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18
Q

Which radical is produced non-enzymatically

A

Hydroxyl radicals

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19
Q

How can hydroxyl radicals be produced

A

Radiation induced homolytic fission or photolytic cleavage

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20
Q

How cna hydroxyl radicals in vivo generate

A

Through a reacation of metal ions such as iron

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21
Q

How many grams of iron are in the body

A

4

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22
Q

What si the recation called that hydroxyl radicals are produced

A

Fenton reacation which is oxidation of a ferrous iron with hydrogen peroxide

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23
Q

What is ferric trivalent ions and ferrous ions

A

Ferrous iron is iron two and ferric iron is iron 3

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24
Q

What happens with the ferric trivalent ions formed in the fenton reacation

A

Need to be re-generated back to ferrous ions whihc happens in a recation with superoxide anion radical

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25
Q

What is a Harber-Weiss recation

A

Combing recations one and two

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26
Q

What does hydroxyl radicals require

A

Superoxide and hydrogen peroxide

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27
Q

What is the reduction potential

A

Measure in volts of the affinity of a substance for electrons as compared with hydrogen

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28
Q

What does high positive reduction potential mean

A

Thermodynamically they are able to oxidize molecules with lower or negative reduction potential by taking electrons from them

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29
Q

What does hydrogen peroxide have

A

lowest recativity and the highest stabality (measured by its half life) and highest intracellular concentration

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30
Q

What is the most reactive and dangerous ROS

A

Hydroxyl radical

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31
Q

What moleculer has intermediate activity

A

Superoxide

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32
Q

What is the dual function of ROS

A

Either be toxic and cause apoptosis or can also cause cell proliferation depending on the concentration

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33
Q

What type of lipid is susceptible to ROS

A

Unsaturtaed lipids

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34
Q

What is lipid preoxidation

A

Oxidative damage to lipids

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35
Q

What is initation for damage to lipid

A

Unstable lipid radical is produced when it recats with ROS to the allylic hydrogen and allylic carbon

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36
Q

What is an allyic carbon

A

Carbon atom bonded to a carbon atom of a double bond

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37
Q

What is the propagation stage

A

The lipid radical recats with the molecular oxygen creating lipid peroxyl radical which recats with another lipid producing a different lipid radical lipid peroxide which initaties the next cycle of lipid oxidation

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38
Q

What is the termination stage

A

Lipid preoxidation is stopped by antioxidants

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39
Q

What happens if antioxidant mechanisms are imparied for lipids

A

The accumulated lipid peroxides exert toxic effects on cells membrane integrity fluditiy permeability change of enzyme activity and more

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40
Q

What is the most common DNA lesion

A

ROS induced oxidation of Guanine to 8-Oxoguanine
8-Oxoguanie can pair with Adenine which is modifying purines and pyrimidines mismatch pair

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41
Q

Can radiation also induce fragmentation

A

YES

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42
Q

What are the negative consequences of ROS induced damages to nucleic acids

A

Point mutations
Changes in gene expression
Single and double strand DNA breaks

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43
Q

What does oxidative damage do to protiens

A

Change in protien structure, function, turnover and loss or occasional gain of acitivity

change in enzyme activity change in ion transport proteolysis autoimmune respone

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44
Q

What does oxidative damage of proteins means

A

ROS-induced modifications individual amino acid and their functional groups

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45
Q

What can thiol groups on cysteine do

A

They react different types of ROS which lead to the formation of sulfoxidation products or intra protien disulfides they cna be dangerous to cells

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46
Q

What intensive oxidative stress do

A

Induce fragmentation of peptide chain and can aggregate due to cross linked recaations and alterd electrical charge and loss their activtiy

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47
Q

What is the glycocalyx coat

A

Surface of cells that are covered with glycans

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48
Q

What does oxidatvie damage to glycans mostly result in

A

Modfication of individual monsaccharides glycan celvaveg fragmentation degradation and accumation of AGE

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49
Q

What can these modifcations to glycagens do

A

Impairment of intracellular contacts (glycocalyx) induction of inflammation and modified functions of extracellular matrix

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50
Q

What are biomarkers of oxidative stress

A

products of oxidative damage

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51
Q

What are some biomarkers include

A

Lipids
DNA
carbohydrates

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52
Q

What are specalized biomarkers

A

Biomarkers relating to specific conditions exist

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53
Q

What are the organelles known to be sources of oxidative stress

A

Plasma membrane
Mitochondria
ER
Peroxisomes
Lysosomes

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54
Q

ROS direct or indirect products

A

BOTH and can function as parcirine signals

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55
Q

What is the major source of superoxide in the body

A

NADPH oxidase which is assembled in the plasma membrane

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56
Q

How many members of the NOX family oxidase

A

7

57
Q

What is NOX2 a key component of

A

THe phagocyte NADPH oxidase

58
Q

What are neutrophils

A

Cells of the inate immunity comprise white blood cells principle phaogcytic cells that migrate to site of injury or infenction recognize engluf and destory the invading pathogens

59
Q

Can neutrophils generate ROS

A

YES which contribute to inactivation of pathogenic bacteria and fungi

60
Q

How are neutrophils morpholgical different

A

oddly shaped nuclei with several loves and densenly stained granules

61
Q

How many transmembrane domains do NOX family members share

A

6

62
Q

What domains bind asymmetrical hemes

A

III and V each contain two histidine

63
Q

What is heme

A

Iron-containing prosthetic group found in enzymes, ETC and oxygen binding pigments like haemoglobin

64
Q

What do the heme groups do

A

confer functionality which can include oxygen carrying, oxygen reduction electron transfer and other process

65
Q

What is the cytoplasmic C-terminal domain in NOX

A

FAD and NADPH binding domain

66
Q

What kind of enzymes are NOX thought to be and what is the process

A

Single electron transporters passing electrons from NADPH to FAD first heme to the second heme and then finally to oxygen to reduce it into superoxide anion radical

67
Q

Which enzymes contain Ca binding domains

A

NOX5 DUOX1 and DUOX2

68
Q

Which domians contain peroxidases-like domain

A

DUOX1 and DUOX2 which gives them the name of dual oxidases

69
Q

What NOX enzymes are similar in size and domains structure to NOX2

A

NOX1, NOX3, NOX4

70
Q

What is NOX2 also known as

A

gp91phox

71
Q

How do NOX1 NOX3 and NOX4 interact with superoxide

A

Electron transfer centers that you need to pass electrons from cytoplasmic NADPH to molecular oxygen which will form superoxide otuside the cell then it will recact to form hydrogen peroxide which is a substrate for peroxidases

72
Q

How does NOX5 interact with superoxide

A

It has a calmodulin like domain for calcium which makes the enzyme activated by calcium to produces super oxidize

73
Q

How does DUOX create Superoxide

A

Build off the NOX5 structure but adds an amino terminus an extra transmembrane a-helix followed by a peroxidases loacalized on the outside of the plasma membrane

74
Q

Can NOX2 generate superoxide itself

A

NO

75
Q

What is the multicomponent complex also called

A

Phagocyte NADPH oxidase

76
Q

What are the five proteins that should be combined to activate gp91phox

A

p22phox p40phox p47phox p67phox and RAC

77
Q

What is flavocytochrom b558

A

p22phox being a hetrodimer with NOX2 (sits in the lipid raft of the plasma membrane)

78
Q

What forms a trimer to actiatre phagocyte

A

p40phox, p67phox, p47phox

79
Q

What activates the pathway of NADPH oxidases

A

Induced by pathogens or inflammatory mediators or by some specefic compounds

80
Q

What does fMLP stand for

A

N-formyl-L-methionyl-L-leucyl_phenylaaline bacteria wall oligopeptides

81
Q

What are the 3 signlaing process nedded for this system

A

Protien phosphorylation lipid metabolsim and GTPase activation

82
Q

What is the process of activation of phagocyte NADPH oxidase

A

Protien kinase requries for phosphorylation p47 which p47 will now pind too p22 lipid metabolsim is then requried to syntehzies PtdlnsP3 which makes change in the lipid compostion which stabalizes the enzymatic complex. You then need GTP to actiave RAC to stablize the enzymatic complex. p67 is also used to convert NADPH to NADP. It can now produce recative oxygen species

83
Q

What is the job of p47

A

Organizer

84
Q

What is the job of the p67

A

It is an activator

85
Q

What is the job of p40

A

It is a second adaptor

86
Q

What are the essential protein binding domains

A

AIR SH3 PRR PB1 tetratricopeptided repeat T and AD

87
Q

What are the lipid binding domains

A

PX and lipid tail of RAC

88
Q

How are p47 p67 and p40 associated with each other

A

Through PRR-SH3 and PB1-PB1 interactions

89
Q

Why do p40/p47/p67 trimer stays in they cytsol

A

Because of auto-inhibitory region AIR p47phox is combined with its SH3 domain which prevents its binding to the PRR domain of p22phox

89
Q

Why is RAC maintained in the cytsol

A

Its lipid tail is masked by the inhibitory protien RhoGDP-dissociation inhibitor which prevents the binding to the membrane

90
Q

What also blocks the phox protiens from binding to the memrabne

A

The binding of SH3 in p47 and by PB1 in p40phox

91
Q

What trigger en block

A

The phosphorylation of p47phox which serve as an organizer the phosphorylation induces a conformational change of p47 which unmasks the SH3 domains so now PRR of p22phox can bind to SH3 pX is now accesible to the membrane

92
Q

How will p67 be activated

A

Should be oriented in a specefic postion which is controled by RAC and p40 through protien protien and protien lipid interacations

93
Q

How do you activate RAC

A

GTP binds to RAC which can induce conformational changes which will promote dissociation of RhoGDI and membrane association through the tail which will now bind to p67 and induce a conformation change allow the activation domain to act on gp91

94
Q

What is Chronic Granulomatous Disease

A

Mutation in the 5 structural genes of the NADPH oxidase expect RACT

95
Q

A mutation in what accounts for the majority of cases in CGD

A

gp91phox then p47 lease being p40

96
Q

What happens with patients who have CGD

A

Fail or are less able to generate superoxide and have a high sensitivity to bacterial and fungal infections

97
Q

What are granulomas

A

Masses of immune cells that form at sites of infection or inflammation and can result in damage to those tissues

98
Q

What causes a more server recation

A

gp91 phox-deficient

99
Q

What are ROS-sensitive indicators

A

Nitroblue Tetrazolium and Dihydrohodamine

100
Q

How can you tell if there is ROS with Nitroblue Tetrazolium

A

It is a yellow transparent color but when reduced it becomes insoluble and turns a deep purple and it shows that there is a production of oxygen radical without it they have CGD

101
Q

What does the DHR test do

A

Measures ROS using flow cytometry uses a flow cytometry machine to detect signals from each cell.

102
Q

What happens if they only see one peak in DHR

A

This means that the patient has CGD because it is only showing the no flourscent cells that are active

103
Q

What happens if you see two peaks in DHR

A

The patient is healthy because the once you have activation NADPH oxidase normal neturophills will then produce superoxide radical and that will oxidize DHR to highly fluorescen molecules

104
Q

Where else can large amount of ROS come from

A

Generated in the mitochondrial as a leakage from the ETC especially from complexes I and III that create superoxide

105
Q

Where is superoxide formed in complex I and complex III

A

Complex I is only within the matrix whereas complex II is released both into the matrix and the mitochondrial space

106
Q

How does the mitochondria protect against ROS

A

Uses antioxidant enzymes

107
Q

What are antioxidant enzymes the mitochondria uses

A

Manganese superoxide dismutatse which converts superoxide to hydrogen peroxide CAT an GPx and Prx3 to inacative or catbolize hydrogen peroxide

108
Q

What can heme serve as

A

ROS in cells

109
Q

What is haemoglobin

A

Produces a constant flux of superoxide and carries oxygen through your body

110
Q

When does Haemoglobin release superoxide

A

Due to electron delocalization between heme divalent iron Fe2 and O2 which can then lead to the formation of Fe3

111
Q

How do the blood cells prevent possible toxic effects

A

They contain high levels of antioxidant enzymes such as peroxiredoxins and superoxide dismutase

112
Q

What happens when human erythrocytes are exposed to hydrogen peroxised

A

produce echinocytes they are small and throny

113
Q

What is hydrogen peroxide

A

Spontatneous or enzymatic

114
Q

How is spontaneous hydrogen peroxide formed

A

Superoxide anion radical due to the dismutation recation can happen slow but still work in cells it is catalyzed by enzymes called superocide dismutases

115
Q

Why is SOD important in antioxidant defense

A

Converts a highly reacative superoxide into less active non radical hydrogen peroxide

116
Q

What are the three types of SOD and where are they located

A

SOD1 (in the cytoplasm) SOD2(in the mitochondria) SOD3 depends on copper zinc or maganese as cofactors

117
Q

How else can hydrogen peroxide be generated

A

Enzymes called oxidases which catalyze an oxidation-reduction reacation molecular oxygen is the receptor

118
Q

What are peroxisomes

A

Generation and scavenging of hydrogen peroxide

119
Q

How can you see peroxisomes

A

With GFP using fluorescence microscopy as mutiple dots in the cytoplasm

120
Q

What do peroxisomes contain

A

Hydrogen peroxide-degrading enzymes and SOD

121
Q

What does Catalsae do

A

Directly incatvaties hydrogen peroxide converting it to water and oxygen

122
Q

What does peroxidase do

A

Utlize hydrogen peroxide as a a substrate for the oxidation of certain organic compounds

123
Q

What does peroxiredoxin 5 do

A

Major scavenger of highly eacative peroxyntirie

124
Q

What happens to peroxisomes when under oxidatve stress

A

Tubular peroxisms they have a higher surface and can accomodate more antioxidant enzymes to prevent oxidative damage

125
Q

What does NAC do

A

Peroxismes wont become tubular because it is preventing oxidative stress

126
Q

What does P450 do

A

Produce ROS as by products it is also involved in metabolic conversion of lipophilic substrates by oxdizing them to become more polar and soluble to become secreted from cells

127
Q

How is ROS produced in cytochrome P450

A

Heme ferrous/ferric states there are two where there are leakeges of a large fraction of the activated oxygen from the enzyme without substrate oxidation

128
Q

What system is P450 apart of

A

monoxygenase system oxidizes a substrate in a complex process involving the electron transfer from NADPH through NADPH-cytochrome reducated to cytochrome P450

129
Q

What does the autoxidation shunt produce

A

Fe Iron (II) superoxide autocidation of the oxy ferrous complex

130
Q

What does the peroxide shunt produce

A

Hydrogen peroxide by the protonation of the peroxyctochrome ferric complexi (III)

131
Q

What is myeloperoxidase

A

Specefic enzyme of neutrophils localized in azurophil granulaes and helps destroy invading pathogens

132
Q

What does MPO catalzye

A

Production of highly recative HOCl in a recation of hydrogen peroxide and cholrine ions

133
Q

Is HOCL powerful

A

Yes more powerful than hydrogen peroxide and shows how moderate ROS can be converted to more aggresive molecules

134
Q

What is peroxynitrite

A

Nitric oxide and superoxide

135
Q

What is NO

A

A free radical generated under phsyiological conditions in all mammal cells called NOS

136
Q

What are the three types of NOS

A

Enothelila neuronal and inducible but function in the same way and produce NO and citrulline from arginine you need oxygen and NADPH as an electron donor

137
Q

What can NO do

A

Vasodilationa and neutrotransmission and antimicrobial and antitumor activties

138
Q

What can NO do

A

Depletion of SH groups
Oxidation
Nitration

Cell Bio (13 decks)

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