protein function - chp6

Question 1

what specific mutation was identified in the alpha subunit of this family’s hemoglobin?

Answer 1

Histidine (His) a hydrophilic amino acid at position 58, was replaced by Leucine (Leu), a hydrophobic amino acid, known as the H58L mutation

Question 2

why was the father benefitting from smoking and asymptomatic for anemia, despite the daughter?

Answer 2

smoking generates a very small amount of CO, and CO is able to bind to hemoglobin in spite of the mutation to Leu58, leaving the entire hemoglobin molecule able to continue picking up oxygen, and not degrading and destroying itself

Question 3

what are the primary functions of Myoglobin and Hemoglobin?

Answer 3

both proteins bind oxygen reversibly to iron in a poryphyrin ring tightly bound to the protein

Question 4

where is hemoglobin primarily found and how much does it contribute to cell weight, as well as it’s main function?

Answer 4

it is the major protein in red blood cells, accounting for approximately 35% of the dry weight of red blood cells. it primarily transports O2 from the lungs and tissues through the circulatory system

Question 5

where is myoglobin concentrated and what is its primary function?

Answer 5

its concentrated in muscles and is a storage depot for O2

Question 6

what prosthetic group is required for both myoglobin and hemoglobin to bind oxygen?

Answer 6

a heme group (specifically a prophyrin ring) that is tightly bound to the protein

Question 7

what is the biochemical cause of Porphyria, also known as vampire disease?

Answer 7

it’s caused by defects in heme synthesis, which leads to an inability to make a porphyrin ring

Question 8

why do individuals with vampire disease avoid sunlight?

Answer 8

in lieu of poryphyrin rings, they have a build up of intermediates that are UV reactive, causing their skin to blister in the sun

Question 9

which enzyme is involved in the porphyrin chemical pathway?

Answer 9

Uroporyphyrinogen decarboxylase (UroD)

Question 10

what is the specific chemical state of iron required for the oxygen to bind to heme?

Answer 10

oxygen only binds to the heme with iron in its reduced (Fe2+) state, which appears as bright red

Question 11

what is heme?

Answer 11

it is the Fe-Porphyrin complex

Question 12

what are the two primary molecular building blocks required for heme synthesis?

Answer 12

an amino acid (glycine) and a citrate cycle intermediate (succinyl-CoA)

Question 13

why can’t iron exist as Fe3+ in heme?

Answer 13

it’s unable to bind to oxygen in this state, as it’s very insoluble and unable to react, and appears as bluish dark brown

Question 14

what is the general structure of the heme group as shown in the diagram?

Answer 14

it is a complex organic ring structure with a central iron (Fe2+) atom coordinated by four nitrogen atoms

Question 15

what is the polypeptide composition of myoglobin?

Answer 15

myoglobin consists of a single polypeptide chain with one heme group, therefore having a tertiary structure (3), binding to only a single oxygen (one heme group)

Question 16

describe the structure of hemoglobin?

Answer 16

hemoglobin is a tetramer consisting of four polypeptides with 2 alpha subunits and 2 beta subunits, able to bind to 4 oxygen at a time (as has 4 heme groups)

Question 17

what is the globin fold?

Answer 17

it is a shared protein fold found in myoglobin and both hemoglobin subunits, containing 8 alpha helices

Question 18

how much sequence identity exists between the amino acids of Myoglobin and Hemoglobin?

Answer 18

less than 20% of the amino acids are identical between myglobin and hemoglobin subunits, despite sharing the same overall fold

Question 19

how many heme groups are present in a single hemoglobin molecule?

Answer 19

four, one for each of the four subunits

Question 20

which specific amino acid residues coordinate the iron in the heme group?

Answer 20

the proximal histidine (His F8) and the distal histidine (His E7)

Question 21

how is the oxygenation of myoglobin and hemoglobin bound?

Answer 21

it is bound through the 6 coordination of Fe2+ with 4 tetrapyrole ring (N), 1 proximal Histidine (R) and 1 O2 (six coordinates in total)

Question 22

what is the role of Distal Histidine in oxygen binding?

Answer 22

it forms a hydrogen bond with the O2 ligand to stabilize the binding

Question 23

which diatomic molecule besides oxygen can bind to the heme group with much higher affinity?

Answer 23

Carbon monoxide (CO)

Question 24

how does the binding affinity of Carbon monoxide compare to that of Oxygen (O2)

Answer 24

CO has a binding affinity for the Fe2+ in hemoglobin that is 200x stronger than that of oxygen (explains carbon monoxide poisoning)

Question 25

what type of coordination geometry is formed when Fe2+ or a metal (M) binds with EDTA?

Answer 25

it forms an octahedral coordination complex

Question 26

what is the function of the EDTA darts (vampire theme) shown in the movie Blade II?

Answer 26

the darts are used to deliver EDTA which acts as a chelating agent to displace or disrupt the metal coordination in the target's system (grabs all the iron from the blood and kills the vampire)

Question 27

how does the structure of the mEDTA-complex compare to heme binding?

Answer 27

while heme uses a porphyrin ring and histidines to coordinate iron, EDTA wraps around the metal ion using its own nitrogens and oxygens to satisfy the octahedral sites

Question 28

what is the physical state of the heme group when oxygen is not bound?

Answer 28

without oxygen, the heme is not planar because the ion is too large, resulting in a pucker, this is called deoxyhemoglobin

Question 29

what happens to the ferrous ion (Fe2+) upon oxygen binding to allow it to move into the porphyrin plane?

Answer 29

the shared electrons make the radius of the ferrous ion slightly smaller, allowing it to move into the plane of the porphyrin ring

Question 30

how does the movement of the iron atom affect the rest of the protein structure?

Answer 30

as the iron moves into the plane, the proximal His is translocated closer to the heme, which pulls on the F helix, leading to more pronounced conformation changes in hemoglobin than myoglobin

Question 31

why are conformation changes upon oxygen binding more pronounced in hemoglobin than myoglobin?

Answer 31

due to hemoglobin's quaternary structure (arrangement of four subunits)

Question 32

what are the two conformational states of hemoglobin?

Answer 32

T state (deoxyhemoglobin) tense state R state (oxyhemoglobin) relaxed state

Question 33

which specific amino acids residues form a hydrogen bond in the T state?

Answer 33

ASP94 on the alpha1 subunit forms a hydrogen bond with Asn102 on the beta2 subunit

Question 34

how does the hydrogen bonding pattern change when hemoglobin transitions to the R state?

Answer 34

the bond between Asp94 and Asn102 is broken - a new hydrogen bond forms between Asp94 and a different site on the B2 subunit - Tyr42 (A1 subunit) participates in hydrogen bonding with Asp99 (B2) in the T state, which changes during the shift

Question 35

what serves as the trigger for the T to R conformational shift?

Answer 35

oxygen binding to the heme iron, which pulls the F-helix and reorganizes the interface between the alpha beta dimers

Question 36

is oxygen binding cooperative in myoglobin, hemoglobin or both?

Answer 36

cooperative oxygen binding occurs in hemoglobin, but not myoglobin

Question 37

how does the affinity of the first oxygen molecule compared to the fourth oxygen molecule binding to the hemoglobin?

Answer 37

the fourth O2 binds with an affinity 100x higher than that of the first O2

Question 38

what is the mechanism behind cooperative binding in hemoglobin?

Answer 38

small structural changes induced by oxygen binding in one region of the hemoglobin leads to large structural changes int the entire hemoglobin molecule, resulting in increased O2 binding for the rest of the subunits, aka positive cooperativety

Question 39

what term described the communication between the distant binding sites in a protein like hemoglobin?

Answer 39

known as the positive allosteric effect (or cooperativity), where the activity at one site alters the activity at another

Question 40

what is the shape of the oxygen binding curve for myoglobin versus hemoglobin?

Answer 40

myoglobin : hyperbolic curve hemoglobin : sigmoidal curve (s-shaped) due to cooperative binding

Question 41

why is hemoglobin a more efficient oxygen transporter than myoglobin?

Answer 41

because its sigmoidal binding curve allows it to be highly saturated in the lungs but release a much larger percentage of its oxygen (60%) at the lower $pO_{2}$ found in tissues, compared to myoglobin which only releases 20%

Question 42

what is the definitions of allosteric control in the context of protein function?

Answer 42

a biological process where the binding of a molecule at one site of a macromolecule (like a protein) causes a functional change at a distant site, altering the macromolecule's activity

Question 43

which two molecules are examples of positive allosteric effects in hemoglobin?

Answer 43

oxygen and carbon monoxide

Question 44

what are the three negative allosteric regulators of hemoglobin?

Answer 44

pH / CO2 and 2,3-biphosphoglycerate

Question 45

what is the metabolic origin of 2,3-BPG?

Answer 45

it is derived from an intermediate of the glycolysis pathway in red blood cells

Question 46

where does 2,3-BPG bind to the hemoglobin molecule?

Answer 46

it binds to the central cavity of the hemoglobin, located between the two beta subunits of hemoglobin

Question 47

how does 2,3-BPG affect the conformational state of hemoglobin?

Answer 47

it traps hemoglobin in the deoxy state (T state)

Question 48

what interactions stabilize 2,3-BPG in the central cavity of hemoglobin?

Answer 48

stabilized mainly by ionic interactions, as the amino acid residues are basic and positively charged, and 2,3-BPG is negative

Question 49

what is the functional consequence of 2,3-BPG binding in red blood cells?

Answer 49

it acts as a negative allosteric regulator, reducing hemoglobin's affinity for oxygen and promoting its release to tissues