CH 4 (L10)

Question 1

Myoglobin

Answer 1

(Mb), Myo refers to muscle, exists as a monomer in muscle of reptiles, birds, and mammals, found in muscle

Question 2

Hemoglobin

Answer 2

(Hb), hemo, heme refers to blood, exists as a tetrameric complex
- heme consists of a tetrapyrrole ring system connected by methene bridges (-CH=)

Question 3

red blood

Answer 3

red color associated with blood due to the heme prosthetic group: protein-bound organic molecule essential for the activity of the protein
–> ferrous iron is bound and forms a complex with 6 ligands, 4 being nitrogen atoms of the protoporphyrin IX

Question 4

sperm whale oxymyoglobin

Answer 4

myoglobin is part of a family proteins called globins
–> myoglobin consists of 8 alpha helices in a bundle –> all-alpha helical bundle –> they are angled to allow the R-groups to interdigitate

Question 5

Myoglobin (Mb)

Answer 5

interior is almost all hydrophobic amino acid residues

• -> the surface is made of both hydrophilic and hydrophobic residues
• –> the heme prosthetic group occupies a hydrophobic cleft stabilized by weak interactions
• –> accessibility to oxygen depends on nearby amino acid side chains

Question 6

Neuroglobin (Ngb)

Answer 6

another globin, recently discovered

• -> preferentially expressed in nervous system tissue
• -> monomer, 150 aa in length, 16kDA, typical globin fold, binds O2, CO, or NO
• provides neuroprotection

Question 7

Cytoglobin (Cygb)

Answer 7

another recently discovered Globin
–> expressed widely in tissues, can bind O2, may be involved in protection during oxidative stress, 1900aa in length (human Cygb)

Question 8

Erythrocytes

Answer 8

erythro-red, cytes- cells
oxygen and hemoglobin carrying the cells of the blood in vertebrates,
each 6-8 um in diameter cell carries ~ 3x10^8 molecules of Hb

Question 9

Hemoglobin (Hb)

Answer 9

• an (α2β2) alpha2beta2 tetramer made of α-globin (7 α helices) and β-globin (8 α helices)
• each globin is similar to myoglobin in structure
• each of the four contains a heme prosthetic group
• 1 α chain interacts extensively with a β chain so sometimes Hb is referred to as an αβ dimer

Question 10

oxygenation

Answer 10

refers to the reversible binding of the oxygen to the heme group

• -> without oxygen , Hb is called deoxyhemoglobin
• -> with oxygen, it is called oxyhemoglobin

Question 11

tissue distribution of vertebrate globins

Answer 11

blood - hemoglobin
muscle - myoglobin
neurons - neuroglobin
all organs - cytoglobin

Question 12

Hb

Answer 12

hydrophobic residues allow heme prosthetic group to be buried inside the hydrophobic cleft

• -> 4 nitrogen help coordinate the iron Fe II
• -> the fifth ligand is an imidazole ring (His-93) called the proximal Histidine
• -> the 6th ligand is molecular oxygen (coordinated to the Fe II and imidazole side-chain of His-64

Question 13

Holding in Place

Answer 13

free heme (in solution) does not reversibly bind oxygen in aqueous solution

• -> Fe2+ is partially oxidized to Fe3+ when oxygne is bound
• -> an electron is temporarily transferred toward the oxygen atom attached to the iron (dioxygen is partially reduced)

Question 14

Reversible binding of oxygen

Answer 14

binding of oxygen depends on

• -> concentration of protein
• -> concentration of oxygen

Question 15

Fractional saturation (Y) of myo/Hb

Answer 15

measured as the amount of which has molecular oxygen bound divided by total (bound + unbound)

Formula:
Y = [MbO2] / [MbO2] + [Mb]

Question 16

Oxygen-binding curves

Answer 16

the curve for myoglobin is hyperbolic (so just one equilibrium constant for binding)
- the curve for hemoglobin is sigmoidal (S-shaped) indicating more than one molecule is binding –> 4 molecular oxygen binds –> these represent environmental pO2 levels

• -> oxygen binds in a positive-cooperative fashion (1 bound will help the 2nd bind which will help the 3rd)
• -> physiologically at 3 this stops as only 1 slot is open per Hb molecule in vivo
• -> notice the pressure at half- saturation (P50)

Question 17

Partial pressure

Answer 17

• is a measurement of the percentage of a give constituent in the gas according to the total pressure
  1 atm - 760 mmHg
• the partial pressure of oxygen (~21% of the atmospheric gases) at 1 atm = 760 x 0.21 = ~160 mm Hg

Question 18

The cooperative binding wiggle

Answer 18

• when the heme iron (red) of hemoglobin is oxygenated, the proximal histidine residue is pulled towards the porphyrin plane
• this disrupts ion pairs that cross-link deoxyHb helping cooperative binding to occur

Question 19

conformation changes

Answer 19

• explain the sigmoidal curve of Hb
• T conformation (tense) resists binding oxygen (low linear line)
• R conformation (relaxed) facilitates binding of oxygen (high curve line)
• -> hemoglobin observed in the middle
• R and T states are in dynamic equilibrium

Question 20

allosteric action

Answer 20

binding of oxygen to Hb is regulated by allosteric interactions

Question 21

allosteric modulator or allosteric effector

Answer 21

binds to a protein and modulates its activity

Question 22

allosteric protein

Answer 22

protein regulated by allosteric effectors

–> substrate/ligand binds when the protein is in R state

Question 23

Allosteric inhibitors

Answer 23

bind their own regulatory sites and bind most avidly to the T state

Question 24

Allosteric modulation

Answer 24

carried out by conformational changes

Question 25

Allosteric proteins

Answer 25

- are in equilibrium when their R states and T states are rapidly interconverting

Question 26

2,3-biphosphate-D-glycerate

Answer 26

- 2,3BPG is an allosteric effector of Hb in mammals - When 2,3BPG is present, the P50 for binding of O2 in Hb goes from 12 to 26 torr - 2,3BPG binds to deoxyHB and stabilizes the T state - What does that mean for binding to oxygen? Lower affinity - In the R state, the binding pocket is too small for 2,3BPG - O2 increases affinity - 2,3BPG decreases affinity - -> Without 2,3BPG, Hb would not release oxygen at the low levels found in tissues - -> 2,3BPG allows oxygen to follow its concentration gradient into tissues where it is needed

Question 27

Enter CO2

Answer 27

- -> OxyHb binding is further regulated by the buffering system in the blood - -> CO2 decreases the affinity of Hb for O2 by lower pH inside red blood cells - -> CO2 is hydrated in RBCs to produce carbonic acid which dissociates to form bicarbonate and a hydronium ion Formula: CO2 +H20 H2CO3 H+ + HCO3-

Question 28

Changes in pH

Answer 28

- Lower pH in the RBCs leads to a change in the environment - -> There are amino acids with ionizable side chains... so what do you think can happen? - -> changes in protonation to several groups in Hb (alpha-amino groups with pKa values around 7-8)

Question 29

Carbamate Adducts

Answer 29

- most CO2 is carried in the blood as bicarbonate ions | - -> some is carried by Hb when CO2 reacts with the N-terminus of Hb to form carbamate adducts

Question 30

The Bohr Effect

Answer 30

These stabilize the deoxyHb conformation. - -> P50 of Hb rises: this is called the Bohr effect - -> What happens to the oxygen that was on the Hb? it gets unloaded more easily

Question 31

Oxygen-hemoglobin Dissociation: Exercise

Answer 31

you exercise heavily --> your mitochondria churn in your tissues --> your muscles use of a lot of oxygen --> you produce more CO2 which needs to be unloaded in lungs --> CO2 causes decrease in pH in RBCs --> Hb-O2 affinity decreases Net effect: more CO2 is unloaded, O2 more easily gets into tissues

Question 32

Bohr effect

Answer 32

low pH (more acidic), high temp, high P of CO2, high 2,3-BPG

Question 33

Immunoglobulins

Answer 33

antibodies (Ig) are part of the vertebrate immune system - -> highly specialized glycoproteins - -> they are highly adapted to binding to portions of foreign molecules to help eliminate them (antigens) - -> they are produced by B-lymphocytes (B cells) --> IgG: most abundant form of immunoglobulin in your blood

Question 34

IgG

Answer 34

most abundant form of immunoglobulin in your blood - 2 heavy chains - 2 light chains

Question 35

immuglobulin fold

Answer 35

- most common motif for the antibody is the Ig Fold domain (110 residues) - 2 antiparallel beta sheet sandwich - the variable region of an antibody is so specific, no water molecules are thought to exist between the antibody and the antigen it binds

Question 36

binding antigen

Answer 36

- hydrogen bonds and electrostatic charges stabilize the binding