KIN217 20-25CH

Oxidative phosphorylation location

Mitochondria

Mitochondria evolutionary endosymbiosis

-Was once and independent organism that was engulfed by another cell

-Has some DNA

Mitochondrial outer membrane

-defines intermembrane space

-permeable to small ions and molecules through mitochondrial porin

Mitochondrial inner membrane

-defines matrix

-folded into cristae

-high protein content

-high amounts of cariolipin

Heart vs liver mitochondrion

Heart mitochondrion have many more cristae allowing for more energy production

Electron transfer potential

Measure of as molecule’s tendency to donate or accept electrons

Negative E0’

-reducing agent that donates electrons

-low e- affinity

Positive E0’

-oxidizing agents that accept electrons

-high e- affinity

Electron movement in the chain

From low E0’ to high E0’

NADH-Q oxidoreductase

-complex I

-accepts 2 e- from NADH and trasfers to ubiquinone

-pumps 4 protons

Succinate-Q reductase

-complex II

-accepts e- from FADH and transfers to ubiquinone

-not a proton pump

Q-cytochrome c oxidoreductase

-complex III

-accepts e- from QH2 and transfers to cytochrome c

-pumps 4 protons

Cytochrome c oxidase

-Complex IV

-accepts 4 e- from cytochrome c and transfers to oxygen

-pumps 4 protons

Respirasome

-complexes of the ETC associated with one another

-mainly complexes I, III, IV

QH2

Reduced form of ubiquinone, carries electrons

Succinate dehydrogenase

-Kerb’s cycle enzyme, reduces FAD to FADH2

-converts succinate to fumarate

coenzyme Q (ubiquinone)

Serves as a shuttle for electrons between complexes I/II, and III

Process of Q-cytochrome c oxidoreductase

-Cyt C can only receive 1e- at a time

-1st QH2 gives Cyt C 1e- (pumps 2 protons)

-2nd QH2 gives 1e- to CYT C and other to make Q

Cytochrome C

-protein with heme group

-carries 1 e- on a heme iron

-transfers electrons from complex III to complex IV

Protons pumped per NADH

-Total of 10

-4 at complex I

-4 at complex III

-4 at complex IV

Protons pumped per FADH2

-Total of 6

-4 at complex III

-2 at complex IV

reactive oxygen species

-ROS

-result of partial reduction of oxygen

-can oxidize other compounds spontaneously

ROS defence

-Superoxide dismutase

-catalase 

-exercise increases the expression of these enzymes

Proton gradient

-the respirasome pumping of electrons results in an unequal distribution of protons

-electrochemical gradient

Proton motive force

-use the energy of the proton gradient to drive ATP synthesis

-protons flow through ATP synthase like a waterwheel

energy of ATP synthesis

-endergonic process

- +30.5kJ/mol

F0 component

-embedded in the inner mitochondrial membrane

-contains the proton channel

F1 component

-protrudes in the mitochondrial matrix

-contains 3 catalytic beta subunits, each in a different form

Connection of F0 and F1 subunits

-gamma subunit

-external column

Beta subunits

contains active sites that generates ATP

Alpha subunits

isolate beta subunits from each other

ATP synthase and shape of mitochondria

-form dimers and cluster together to form curvatures in the cristae of the mitochondria

-clustering stabilizes rotational forces, increasing efficiency 

O (open) form

nuleotides can bind to or be released from the beta subunit

L(loose form)

nucleotides are trapped in the beta subunit

T(tight) form

ATP is synthesized from ADP and Pi

Subunit a

-2 half channels for proton flow in F0

-one opens into intermembrane space, one into matrix

Ring of c subunits spinning

-glutamate residue at spot on c subunit is exposed to each half channel of subunit a

-protons bind to gluamate to form glutamic acid

-subunit c with no charge can then move into nonpolar region of phospholipids

What dictates direction of spin

-proton gradient

-entry of protons into half channel

protons and ATP

-nned 4 protons per ATP

-8 protons per spin

-3 ATP per spin

-1 proton for substrate availability/ATP export

ATP-ADP translocase enzyme

-antiporter

-ATP export is coupled to ADP import

Pi into matrix

-phosphate carrier

- -OH antiporter 

ATP synthasome

ATP synthase + ATP-ADP translocase + phosphate carrier

NADH from glycolysis

cytoplasmic NADH needs to get into matrix before being used by ETC

glycerol-3-phosphate shuttle

-prominent in muscle

-cytoplasmic NADH transfers e- to DHAP to form glycerol-3-phosphate

-G-3-P transfers e- to FAD in mitochondrial G-3-P dehydrogenase

-FADH2 transfers e- to Q

glycerol-3-phosphate shuttle control

shuttle functions regardless of matrix [NADH]

malate-asparate shuttle

-prominent in heart and liver

-cytoplasmic NADH transfers e- to oxaloacetate to form malate

-malate transfers e- to NAD+ in matrix to go back to oxaloacetate

High martix NADH can inhibit shuttle

malate-aspartate shuttle control

ATP from matrix NADH (from ETC)

2.5

ATP from FADH2 (from ETC)

1.5 (FADH2 doesn’t pump as many protons, not in complex I)

ATP from cytoplasmic NADH

-G-3-P dehydrogenase transport 

~1.5 ATP

-malate-aspartate transport ~2.5 ATP

total ATP per molecule of glucose

30-32 ATP

control of oxidative phosphorylation

-dictated by [ADP]

-electrons do not flow from fuel to O2 unless there is a need to make ATP

proton gradient making heat

-uncoupling proton gradient from ATP synthesis

-protons flow through uncoupling proteins

-energy not captured chemically and released as heat

brown fat

-specializes in uncoupling 

-high in hibernating animals

-in humans high in infants, higher in females

-can be increased with cold exposure

thermogenesis and weight loss

-increased heat production and less ATP prdocution

-not efficient metabolism

-dangerous

Glycogen

-a highly branched homopolymer of glucose present in all tisues

-formed and stored in the cytoplasm

liver glycogen

-10% by weight

-maintains blood glucose when fasting

muscle glycogen

-2% by weight but larger store than liver due to higher muscle mass

-provides glucose for sudden strenuous exercise

glycogen stucture

-straight chains have alpha-1,4,-glycosidic bonds

-branch chains(every tenth residue) have alpha-1,6-glycosidic bonds

-non reducing ends (OH) on exterior

glycogenin

protein at the core of glycogen

glycogen breakdown steps

-degraded glycogen

-remodel glycogen

-convert breakdown product into usable product

use of G-6-P after release

-enter glycolysis

-in liver it can be converted into free glucose

-processed in the pentose phosphate pathway 

cleaving glycogen

-glycogen phosphorylase degrades glycogen from the nonreducing ends

-catalyzes a reaction that yields glucose 1-phosphate

remodeling glycogen problems

-glycogen phosphorylase cannot cleave the 4 glucose residues near branch points

-glycogen phosphorylase can only cleave alpha-1,4-glycosidic bonds

remodeling glycogen solutions

-bifunctional debranching enzyme

-transferase and alpha1,6-glucosidase activity

debranching transferase activity

-shifts small oligosaccarchides near the branch point to a nearby chain

-exchange of alpha-1,4-glycosidic bonds

-shifted glucose moieties become accessible to phosphorylase

debranching alpha-1,6-glucosidase activity

-cleaves alpha-1,6 bond at the branch point using H2O

-releases a free glucose (no phosphate)

making glucose-6-phosphate

-phosphoglucomutase

-enzyme has a phosphorylated serine residue, adds phosphate onto C-6

-removes phosphate from C-1

glycogen phosphorylase

-key enzyme for regulating glycogenolysis

-has a more active ‘a’ form and less active ‘b’ form

-active ‘a’ form has phosphorylated serine

glycogen phosphorylase a and b

-both have R and T state

-in the ‘a’ form, R state is favoured

-in the ‘b’ form, T state is favours

muscle glycogen phosphorylase

-default is the ‘b’ form in T state

-when energy is needed shifts to R state by binding of AMP

-in high energy shifts to T state with binding or ATP or glucose-6-phosphate

epinephrine and muscle glycogen phosphorylase

-phosphorylates enzyme to active a form

-active regardless of ATP, AMP or glucose-6-phosphate

liver glycogen phosphorylase

-default is the ‘a’ form in R state

-glucose as a negative feedback inhibitor

hormonal regulation of liver glycogen phosphorylase

-glucagon/epinephrine activates

-insulin deactivates

phosphorylating glycogen phosphorylase

-adding phosphate shifts from ‘b’ state to ‘a’ state

-maximally active when phosphorylated and bound to calcium

glycogen and fatigue

-fatigue is associated with depletion of glycogen stores

-low glycogen may result in increased ADP

UDP glucose pyrophosphorylase

-substrates are UTP and G-1-P

-products are UDP-glucose and pyrophosphate

-reversible

pyrophosphates

-hydrolyzes pyrophosphate to make 2Pi

-irreversible

glycogen synthase

-key regulatory enzyme

-transfers glucose from UDP glucose to C-4 of the terminal residue of the glycogen chain

-can only add glucose to chain of 4 or more residues

glycogenin

-glycogen priming enzyme

-makes alpha-1,4 chain

-remains attached to glucose via tyrosine residue

branching enzyme

-makes alpha-1,6 linkage

-breaks off alpha-1,4 linkage with ~7 glucose

branching enzyme specifics

-chain tha twas broken must have at least 11 glucose

-new alpha-1,6 linkage is at least 4 residue inwards

glycogen synthase and regulation

-active in unphosphorylated ‘a’ form

-inactive in phophorylated ‘b’ form

-G-6-P stabilizes active R state of b form

reciprocal regulation of glycogen breakdown and synthesis

-glycogen synthesis is inhibited by glucagon and epinephrine

-activated with insulin

-same signalling pathways that stimulate glycogen breakdown

PP1

-protein phosphatase 1

-shifts glycogen metabolism to synthesis

-removes phosphates from glycogen synthase b

epinephrine/glucagon and PP1

-need glucose signal

-activates pkA, phosphorylates regulatory subunit Gm

-allows inhibitor to bind

insulin and PP1

-store glucose signal

-phosphorylates and inactivates glycogen synthase kinase

-PP1 can dephosphorylate glycogen synthase to activate

type 1 diabetes

-autoimmune attack on beta cells

-no insulin production

-poor uptake of glucose, high blood glucose

type 2 diabetes

-more common

-insulin resistance

-glucose uptake impaired, high blood glucose

-insulin still inhibits hormone sensitive lipase in adipose so low risk of acidosis

natural selection and type 2 diabetes

-polygenic

-people with ancestry that had a history of surviving starvation/ famine are at higher risk