SYLLABUS 5: Mitochondrial Carriers and Substrate Shuttles: the Pentose Phosphate Pathway

Question 1

purpose of mitochondrial carriers?

Answer 1

transport metabolic substrates into and out of the mitochondria, b/c of the limited permeability of the mitochondrial membrane

Question 2

carrier 1, phosphate, action

Answer 2

phosphate carrier:

exchanges Pi with OH

Question 3

carrier 2, dicarboxylate, action

Answer 3

dicarboxylate carrier:

exchanges Pi or malate or succinate for each other

Question 4

carrier 3, tricarboxylate, action

Answer 4

tricarboxylate carrier:

exchanges citrate, isocitrate, malate, or PEP for each other

Question 5

carrier 4, aKg, action

Answer 5

aKg carrier:

exchanges aKg for malate

Question 6

carrier 5, pyruvate, action

Answer 6

pyruvate carrier:

exchanges pyruvate for OH or ketone bodies

Question 7

carrier 6, glutamate, action

Answer 7

glutamate carrier:

exchanges gultamate for OH

Question 8

carrier 7, aspartate, action

Answer 8

aspartate carrier:

exchanges aspartate for glutamate

Question 9

carrier 8, adenine nucleotide, action

Answer 9

adenine nucleotide carrier:

exchanges ADP for ATP

Question 10

which carriers can malate exchange on?

Answer 10

2: dicarboxylate carrier
3: tricarobxylate carrier
4: aKg carrier

Question 11

what kind of exchange do carriers catalyze

Answer 11

1:1 exchange w/ their partners

Question 12

which carriers funciton in oxidative phosphorylation?

Answer 12

carriers 1 and 8

carrier 1: exhanges Pi with OH - provides phosphate

carrier 8: exchanges ADP for ATP - provides ADp or ATP

Question 13

what is the purpose of mito shuttles?

Answer 13

compounds that can’t enter or leave mito b/c they lack carriers are directly transported by substrate shuttle mechanisms

Question 14

what happens if NADH produced in G3PDH of glycolysis isn’t reoxidized to NAD+?

Answer 14

glycolysis stops because of lack fo NAD+

however, NADH cannot enter or leave mito

thus reducing equivalents of ANDH (electrons, hydrogens) are transported into mito by shuttles

Question 15

specific functions of shuttles?

Answer 15

1) transport reducing equivalents from NADH or NADPH into or out of the mito
2) provide acetyl CoA for fatty acid or cholesterol synthesis
3) provide carbon intermediates for gluconeogenesis

Question 16

what shuttles translocate the reducing equivalents of NADH into the mito?

Answer 16

1) a-glycerophosphate shuttle
2) malate-asparata shuttle

Question 17

what is the purpose of the alpha-glycerophosphate shuttle?

Answer 17

carries 2 electrons of NADH, aka reducing equivalents, from the cytosol into the mitochondrial matrix, using cytosolic glycerophosphate dehydrogenase

Question 18

how does the a-glycerophosphate shuttle work?

Answer 18

DHAP reacts with NADH [both from glycolysis] to produce a-GP [glycerol 3 phosphate] + NAD⁺; catalyzed by cytosolic a-GPDH

NAD+ is thus reoxidized

now, must regenerate DHAP and do something w/ a-GP

a-GP reacts w/ mitochondrial a-GPDH, on the outer surface of the mito inner membrane to regenerate DHAP

mito a-GPDH is linked w/ FAD, so FADH₂ is also produced

FADH2 reacts w/ Q of the ETC to make QH2 which carries the H’s to the ETC

Question 19

what is the movement of reducing equivalents in the a-glycerophosphate shuttle

Answer 19

cytosolic NADH’s reducing equivalents were transferred to DHAP to produce a-GP

a-GP gave those reducing equivalents to mito FAD to produce FADH​2

FADH2 gives those H’s to the respiratory chain

Question 20

do DHAP or a-GP enter the mito during the a-glycerophosphate shuttle?

Answer 20

no, because they don’t have carriers

Question 21

what’s the purpose of the malate-aspartate shuttle?

Answer 21

to reoxidize the NADH from glycolysis, in the cytosol, or to regenerate OAA

Question 22

what is the process of the malata-aspartate shuttle

Answer 22

IN CYTO:

NADH from glycolysis transfers H’s to OAA

OAA is reduced by Malate by cytosolic malate DH

Malate enters Mito via carriers 2, 3, 4

IN MITO:

Malate reacts w/ Mito Malate DH, becomes OAA + Mitochondrial NADH

These H’s from NADH will -> e- transport chain

OAA undergoes transamination reaction w/ glutamate to make aspartate; glutamate becomes a-KG

Aspartate leaves mito via Carrier 7, a-Kg leaves via Carrier 4

IN CYTO:

Reverse transamination reaction occurs: a-Kg reacts w/ Aspartate, regenerate Glutamate from a-Kg and OAA from Aspartate

Question 23

purpose of the isocitrate shuttle?

Answer 23

transfer the reducing equivalents of mitochondrial NADPH to cytosolic NADP to make cytosolic NADPH

**transhydrogenase **enzyme does this

need to do this b/c NADPH is needed for biosynthetic reactiosn that occur largely in the cytosol

Question 24

how does the isocitrate shuttle work?

Answer 24

NADPH in the mitochondrial matrix reacts with a-KG and CO2 and the mitochondrial isocitrate DH to produce NADP+ and isocitrate

isocitrate leaves the mito via carrier 3

in the cytosol, cytosolic isocitrate DH acts on isocitrate and NADP+ to make NADPH and a-KG and CO2

a-KG returns to the mito via carrier 4

Question 25

where does the pentose phosphate pathway occur?

Answer 25

cytosol of certain tisses - the liver, adipose, mamary gland, steroidogenic tissues like the adrenals, and RBCs

Question 26

where is the PPP inactive / low activity?

Answer 26

brain, uscle, heart

Question 27

what is the PPP purpose?

Answer 27

utilizes glucose for: 1) production of NADPH -\> synthesis of cholesterol, fatty acids, steroids, detoxification reactions by cytochrome P450 system 2) production of ribose -\> produce ntds for RNA and DNA synthesis; NAD, FAD, CoASH 3) reduction of oxidative stress

Question 28

how does NADP / NADPH regulate the PPP?

Answer 28

NADP availability is limiting to the PPP: need it for the 1st 2 steps of the pathway

Question 29

how does insulin regulate the PPP?

Answer 29

insulin stimulates the 1st enzyme of the PPP, G6PDH

Question 30

what is different about the PPP for a tissue that's proliferating vs. not proliferating?

Answer 30

**a tissue that is not proliferating - adipose, mammary, RBC** - continues the PPP once have made 2 NADPH, 1 CO2, and Ribose 5 P because it can **convert that Ribose 5 P into NADPHs ** a tissue that needs both NADPH (for BIOSYNTHESIS) and Ribose-5-P, for RNA and DNA synthesis, can stop the PPP at the Ribose 5 P point

Question 31

how does a cell that isn't doing DNA/RNA synthesis proceed on the PPP?

Answer 31

Rearrangement reactions: Convert Ribose 5P back to G6P to NADPH 3 ribulose 5p -\> 2 fructose 6P + 1 Glyceraldehyde 3P Fructose 6P -\> G6P via isomerase Glyceraldehyde 3P = 0.5 G6P, via gluconeogenesis **THUS, have regenerated 2.5 G6P by rearrangement reactions**

Question 32

what regulates the PPP?

Answer 32

availabilty of G6P and the need for NADPH; NADP availability is crucial for the 2 first steps of the PPP

Question 33

why does the RBC have a PPP if it doesn't divide or synthesize much?

Answer 33

for hemoglobin to bind O2 and take to our tissues, the iron of heme must be in the **ferrous **redox state (Fe2+) this **oxyhemoglobin** can be in resonance where an electron from the Fe2+ is given to the O2, making **peroxyhemoglobin** this **peroxyhemoglobin** can dissociate to **methemoglobin and free O2**, the **superoxide anion radical ** this radical can spontaneously form **peroxide** (hydrogen peroxide at pH=7) H2O2, H peroxide, is a **powerful oxidant that must be rapidly removed to avoid RBC damage ** in order to remove H2O2, the RBC needs to use **catalase and glutathione peroxidase** to **remove Hydrogen Peroxide from our cells. This requires NADPH, **and need a PPP to get that NADPH

Question 34

what happens to peroxyhemoglobin to form peroxide?

Answer 34

3% of peroxyhemoglobin dissociates to methemoglobin and free superoxide anion radical, which can spontaneously form peroxide (hydrogen peroxide at pH=7) or superoxide dismutases catalyze the conversion of superoxide to H2O2 H2O2 is a powerful oxidant that must be removed

Question 35

what deals w/ H2O2 if it is formed in the RBC?

Answer 35

1) catalase, a heme-containing enzyme, decomposes H2O2 to O2 + H20 2) glutathion peroxidase uses glutathione (GSH) to catalyze reaction, via glutathione peroxidase: H2O2+ 2GSH -\> 2H20 + GSSG this GSSG must be converted back to GSH, via glutahione reducase: GSSG + 2NADPH -\> 2GSH + 2NADP+

Question 36

what is GSH?

Answer 36

glutathione the most important antioxidant in cells; the most abundant biochemical in cells it interacts w/ the reactive oxygen species, i.e. peroxide, via glutathion peroxidase reaction to produce H20 and GSSG, gets rid of the active oxidant

Question 37

what is the most common inborn error of metabolism? what does it cause?

Answer 37

G6PDH deficiency high prevalence among African Americans and Mediterranean area ppl deficiency causes decreased G6PDH activity; severity depends on the mutation, and if individual is exposed to oxidant stress from drugs or foods like fava beans or has severe infection whereby immune system produces oxygen radicals decreased G6PDH activity -\> oxidative damage leading to hemolysis of the RBC and hemolytic anemia **_RBC need G6PDH for both the PPP and glycolysis_**