Bioenergetics 2

Question 1

Electral transfer goes from high/low potential to high/low potential

Answer 1

low -> high. Has to be in close proximity.

Question 2

What is max distance for redox reaction?

Answer 2

25 ängström. Electrons are transferred via sp-orbitals

Question 3

Where do electrons and protons go in NADH dehydr?

Answer 3

Reduced substrate = oxidized by the enzyme. 2 Electrons come into the enzyme, along with 2 protons. Due to transfer of 2 electrons in the enzyme, redox free energy is used for a confirmational change in the enzyme that opens up the proton pumps: protons are moved to the outside. PMF.
Protons go out, electrons stay in

Question 4

Where do electrons/protons go with BC-1 complex?

Answer 4

Oxidation step is at the positive side of the membrane (plasmic space). Reduced substrate = oxidized by the enzyme, releasing 2 electr. And 2 protons. 2 protons stay outside but the 2 electrons move inwards. Bc of special device in these types of enzymes that facilitates electron transport (there are metals). PMF.
Electrons move in, protons stay out

Question 5

Where do electrons/protons go with aa3-type oxidase?

Answer 5

Sort of combination of the 2: Also reduced substrate outside, protons are released outside, electrons move inwards. Not completely to negative site, but somewhere halfway (reaction pocket is somewhere half in the membrane, also composed of metals). Meaning: not full charge separation per electron, but half charge separation. Protons come from the inside, also until half the enzyme. Every proton also contributes to half a charge separation.
Electrons move half in from outside, protons move half in from inside (and stay out from outside)

Question 6

Magnitude of membrane potential (buildup of PMF) depends on:

Answer 6

1) Faraday constant
2) Gas constant
3) Temperature (Kelvin)
4) pH

Question 7

What are neutral enzymes and why is there no charge seperation?

Answer 7

Neutral enzymes:
All chemistry takes place on one side of the membrane
Substrate gets oxidized, releases 2 electrons, 2 protons stay outside. Physically separated, but not separated by the membrane.
No charge separation.

Question 8

What is in the soluble domain of the NADH dehydrogenase?

Answer 8

Yellow dots = In soluble domain of NADH dehydrogenase = 2-iron 2-sulfur clusters. Make up a pathway for transfer to quinone.

Question 9

What subunits does cyt-bc1 have and how are they characterized?

Answer 9

Subunit cyt-b has 2 irons. 2nd subunit = riske subunit = periplasmic with risk iron rulfur cluster (metal). 3rd subunit = cyt-c1 subunit , has a heme iron

Question 10

Cyt-c soluble has..

Answer 10

1 heme iron

Question 11

aa3-type oxidase is characterized by what groups?

Answer 11

aa3-type oxidase: copper electron site that accept the electrons. Transferred to reaction site half in the membrane, composed of haem iron and copper iron.

Question 12

What are the PMF generating ways of the enzymes?

Answer 12

PMF generating ways: Proton pump, Q-cycle & proton pump + opposite electron & proton movements

Question 13

How many charge seperations do you have in total when 4 electrons are moved from NADH to oxygen?

Answer 13

20. NADH dehydr = 8q/4e
    Cyt-bc1 complex = 4q/4e
    aa3-type oxidase = 8q/4e

Question 14

What part of NADH accepts electrons? What do the iron-4-sulfur clusters do?

Answer 14

FMN (flavin mono nucleotide)
Ion-sulfur clusters:
4 characteristic cysteine residues that deliver the sulfur ion that is responsible for ligating each of the 4 iron atoms in this cube. (?)
Involved in 1-electron transfer reactions from FMNH2 to quinone

Question 15

What is one of the most important features of the proton pump?

Answer 15

One of the most important features of proton pump in system 1 = they are connected by a helix.
2 connected helices between the 4 proton pumps. Helix is moved (confirmational change) by free redox energy-> release of protons.

Question 16

What do all proton pumps have?

Answer 16

All proton pumps are evolutionarily related. All have Lys, Glu, prolines reserved. Prolines are responsible for knicking the alpha helix of the membrane. Glu: proton-guiding amino-acid residues.
Lysine: positively charged at neutral pH (7). Not charged at higher pH
Glutamate: pH 3 = neutrall pH7 = negatively charged

Question 17

Are quinol/quinone proteins?

Answer 17

No, molecules

Question 18

What does a ubiquinol look like?

Answer 18

Redox active head, isoprene unit, polyprenylated tail (hydrophobic, can move in the membrane)

Question 19

Cyt bc-1 complex has lp and hp hemes where?

Answer 19

Low potential heme at the outside of the membrane (bl), high potential at the relative inside of the membrane (bh).

Question 20

Where are hemes b and c found and how are they bound?

Answer 20

HEme b = in b subunit of cytbc1, ligated via histidine residues
Heme c = in c subunit, covalently bound via cysteine residues

Question 21

What does the riske protein do in cyt bc 1?

Answer 21

Riske [2fe-2S] cluster: transfers electrons from quinol to heme c1 one at the time.
The Rieske protein moves in cycles from a c1-state to an intermediate state. Every time an electron moves into the enzyme, confirmational change.

Question 22

What characterizes the C1 state?

Answer 22

heme c1 = in close proximity with risk iron sulfur, electron transfer allowance.

Question 23

What happens at the same time that Transfer of the electron into the riske iron sulfur induces the hinge mechanism?

Answer 23

• At the same time: Due to this first oxidation step it induces an additional confirmational change, such that the head of the ubiquinone (bc one electron is already transferred to cyt bc1 complex) will move to cyt b. The other electron will use this pathway to heme bl.

Question 24

When does charge seperation occur during quinol oxidation at the site of the bc1 complex?

Answer 24

When the second electron is transfered from the low potential heme to the high potential heme, which is further away from the enzyme and results in charge seperation from th eproton

Question 25

aa3 type cytochrome c oxidase has .. subunits. Subunit .. has a catalytic site with conserved histidine residues which provide ligation. Electron acceptance site = subunit .. Accepts electron from the soluble cyt C

Answer 25

2, 1 , 2

Question 26

What types of proton channels can be found in the aa3-type oxidase?

Answer 26

K-channel. Transfer from inside to halfway.

D-channel. Pathway overlaps with K-channel, important for proton pumping from inside to outside

Question 27

AA3-type oxidase: electron movement?

Answer 27

electron: -> copperA -> low spin heme -> end up at reaction center: high spin heme. Oxygen is already there, but has to be double reduced (4e) before it can bind.

Question 28

AA3-type oxidase: proton movement and water formation?

Answer 28

. 4 Protons move via the K channel also to the reaction center, 2 water molecules are formed and result of electron movement is result in confirmational change, opens up the proton pump (other channel), 4 protons are moved from the inside of the membrane to the outside.

Question 29

What are movements that contribute to the PMF in aa3-type oxidase?

Answer 29

1) Electrons move from relative outside to reaction center, protons still stay outside. This contributes to PMF.
2) Chemical protons move from the inside to the outside. This also contributes to PMF.

Question 30

What type of enzymes have the same ancestor

Answer 30

Ba3, cbb3 and aa3 type oxidases have the same ancestor (heme-copper oxidase family)

Question 31

For cbb3, aa3 and ba3 name the next characterizations:
electron donor
affinity for oxygen
proton pump

Answer 31

cbb3 electron donor = cyt c
aa3 = cyt c
ba3 = quinol

cbb3 = high oxygen affinity
aa3 = low
ba3  = low

all have proton pumps