MEDS2003 Module 1: Metabolism

Metabolism

All of the chemical reactions that happen in the body

Catabolic pathway

Breaking down molecules, can make ATP or extract H/e- for ETC

Anabolic pathway

Building larger molecules, requires energy (like ATP or reducing power from NADPH, NADH)

Substrate level phosphorylation

Phosphorylating ADP to ATP directly from a substrate, like anaerobic glycolysis

Energy charge

[ATP] + 0.5 [ADP] / [ATP] + [ADP] + [AMP] (AMP has most impact)

Cell response to energy charge

Low charge > make more ATP, high charge > use more ATP

Kinase

Enzyme that catalyses a phosphorylation reaction

Phosphatase

Enzyme that catalyses a dephosphorylation reaction

Phosphorylase

Enzyme that catalyses a phosphorolysis reaction (e.g. glycogenolysis)

Synthase

Catalyse condensation reactions which don't need nTP

Synthetase

Catalyse condensation reactions which need nTP

Dehydrogenases

Catalyse redox reactions, usually used NAD+/FAD as cofactors, named after the substrate being oxidised

NAD+

Nicotinamide adenine dinucleotide, an oxidant that likes to turn alcohol into ketone, becomes NADH + H+, carries 1 H and 1 e

FAD

Flavin adenine dinucleotide, an oxidant that likes to turn C—C into C=C, becomes FADH2, carries 2 H and 2 e

Coenzyme A

Carrier of acyl groups, doesn't diffuse across membrane, used to trap metabolites

General Fuel Oxidation Strategy

1. Strip H/e, break fuel into 2 carbon chunks, 2. Krebs cycle rips H/e from acetate, complete oxidation of carbon to CO2, 3. ETC, pump protons out of mitochondria, turn O2 into H2O, 4. Proton gradient spins ATP synthase to make ATP

7 Key Metabolism Concepts

1. H/e carriers are in short supply, 2. ADP is in short supply, 3. ATP is really stable, 4. Inner mitochondrial membrane is proton impermeable, 5. Protons only enter matrix if making ATP, 6. If proton gradient is high, proton pumps don't work, 7. If one thing stops, they all do

Fatty Acid

Carbon chain with COOH, mainly fully reduced, stored as a triglyceride, energy dense, can't be used in brain

Beta-Oxidation

Repeated breaking of fatty acid chains into Acetyl-CoA chunks, transported into mitochondria w/carnitine, cut off one 2C chunk w/ 1 FAD and 1 NAD+

Blood FA

Loosely travel with albumin, can diffuse into cell, trapped by Acyl-CoA

Cytoplasm trapping of Fatty Acids

Fatty Acyl-CoA, by fatty acyl CoA synthetase (ATP AMP), 'activates it'

Carnitine

Molecule that binds to fatty acyl-CoA to transport it into mitochondria, travels out of mitochondria to be recycled

CAT-1 and CAT-1

Carnitine acyl-transferases, 1 is cytosolic (puts carnitine on), 2 is mitochondrial (takes carnitine off)

Beta-Oxidation Steps

FAD acts to form a double bond, hydration makes OH, NAD+ acts on OH, new CoA cleaves, leaving acetyl-CoA + FADH2 + NADH

Beta-Oxidation Results

With Cn chain, n/2 acetyl CoA, and (n/2 - 1) FADH2 and NADH

Glucose

Reasonably reduced, stored as glycogen, low stored, brain takes priority of it

Glycolysis

Glucose oxidation, cytosolic, all tissues, anaerobic, fast, 2 ATP, 2 Pyruvate, 2 NADH made

Glucose Transporters

GLUT-1 in all cells, GLUT-2 liver and pancreas, GLUT-4 muscle and adipose (insulin sensitive)

Glucose trapping

Glucose trapped in a cell as G6P, by hexokinase, uses 1 ATP

G6P

Glucose 6-phosphate, made by hexokinase

Glycolysis Investment Phase

G6P becomes F6P, which becomes fructose 1,6-bisphosphate, which can be split into two 3-carbon sugar phosphates (uses 2 ATP)

Glycolysis Payoff Phase

2 Glyceraldehyde 3-phosphates, get oxidised with NAD to add another P, get 2 ATP, rearrange molecule, get 2 ATP, now have 2 pyruvates

Overall Glycolysis Yield

Two ATP, two pyruvate, two NADH

Pyruvate becoming acetyl-CoA

Via pyruvate dehydrogenase, create acetyl-CoA, CO2 and NADH

Reduction of pyruvate

Pyruvate becomes lactate, regenerates NAD+

Phosphofructokinase

PFK, uses 1 ATP to add phosphate to fructose 6-phosphate

Protein

A rare fuel source, last ditch resort, inefficient as ammonia needs to be removed

DNP

Dinitrophenol, metabolic uncoupler, allows for proton gradient to dissipate not through ATP synthase, weak acid, hydrophobic even when charged

Coupling

Rate of ATP synthesis exactly matches rate of ATP use, 1kg ATP/kg body weight a day of ATP used, if anything slows, it all slows

Muscle Action

Uses ATP to contract

Type I Muscle

Red, slow, many mitochondria, good blood supply

Type IIb Muscle

White, fast, few mitochondria, poor blood supply

When beginning exercise

Increase in ATP consumption, so quicker ATP production, first fuel is blood glucose, so glucagon increase, insulin decrease

ATP Concentration

5 mM. If < 3 mM, then cells die

BGL Homeostasis

Insulin decreases BGL, glucagon increases BGL

Glucagon effect on WAT

Causes release of fatty acids into blood

Continued exercise

FA-CoA becomes main source of Acetyl-CoA, and Acetyl-CoA negative feedback by inhibiting PDH so BG only becomes lactate, gluconeogenesis in liver

Moderate exercise

Fatty-acid metabolism reaches max, PDH no longer inhibited, pyruvate from BG is oxidised, now using FA and glucose

Strenuous exercise

Fatty acids and glucose are maximum, so now glycogen store in muscle is used to make more glucose

Very strenuous exercise

Fatty acids, glucose, glycose is maximum, glycogen now undergoes substrate level phosphorylation, quick and inefficient

Sprinting

Type IIb muscles, poor mitochondria so can't oxidative phosphorylation, only anaerobic, cytosolic glycolysis, quick ATP but a lot of lactates build up

Creatine Phosphate

in equilibrium, CP + ADP C + ATP, can supply for 5 seconds

Krebs Cycle

All Acetyl-CoA go here, fully oxidise them to CO2, produce NADH, FADH2, uses oxaloacetate as a 4 carbon carrier

Krebs Cycle reactions

6 carbon citrate, make NADH to remove 1 CO2, do that again, make 1 GTP, by rearranging back to oxaloacetate, produce FADH2 and NADH

Krebs Cycle Economics

2 C in, 2 C out, makes 3 NADH (2.5 ATP), 1 FADH2 (1.5 ATP), GTP (so indirectly 10 ATP)

Uncoupling

A hole in the mitochondrial membrane, the proton gradient can dissipate without making ATP, e.g. DNP, which allows for instant NAD and NADH regeneration, massive fuel consumption, but no ATP production

UCP-1

Uncoupling protein-1 (thermogenin), a natural uncoupler, found in brown adipose tissue, used to generate heat, high in neonates

ETC

Electron transport chain, series of pumps that use NADH and FADH2 to move H+ out of the mitochondria, and turns O2 into H2O

NADH ETC Path

I, Q, III, IV

FADH2 ETC Path

II, Q, III, IV

ETC Complexes

I, II, Q, III, IV, made up of structural and prosthetic group, arranged so H+ expelling reactions are on the outside, H+ using reactions are on the inside

NAD/H Spectrometry

NAD+ absorbs at 270 nm, NADH absorbs at 340 nm

ETC Complex I

NADH acts to pump 4H+, provides things for Q

ETC Complex II

FAD is stuck inside, provides things to Q

ETC Complex III

4H+ pumped

ETC Complex IV

2H+ pumped, whilst making water using Oxygen

UQ

Ubiquinone, very hydrophobic, stored in the Q pool

ETC Q pool

Store of UQ, within the inner mitochondrial membrane

Cytochrome C

Picks up e- from III and delivers to IV, uses iron in porphyrin rings or iron-sulphur, iron changes from 2+ to 3+ and back

Proton Pump Mechanism

Hydrogen carriers give only e- to e- carrier, H+ is released, and e- carrier give e- and H to a hydrogen carrier (H+ enters from inside, and leaves outside)

NADH Yield in ETC

10 H+

FADH2 Yield in ETC

6 H+

Outer mitochondrial membrane

Has holes in it, not important to consider

Proton motive force

Electrochemical gradient that drives ATP synthase, it's made by the ETC

Glycerol 3-Phosphate Shuttle

Method of getting NADH's reducing power to ETC complex II (less yield as it bypasses complex I)

Malate Aspartate Shuttle

Malate can enter cell (carries H from NADH), gives H back to NAD inside mitochondria, regenerated with two linked cycles (Malate>Oxaloacetate>Ketoglutarate) and (Glutamate>Aspartate)

Cytosolic NADH Transport Mechanism

Glycerol 3-Phosphate Shuttle (less efficient), or Malate Aspartate Shuttle

Routes to Q Pool

From I, from II, from beta-oxidation making FAD, from glycerol 3-P shuttle

Free Radical Formation

Can form in UQ pool if there is a traffic jam, less likely to form if III is vacant

ATP Synthase

A protein complex which uses H+ gradient like water spinning a water wheel, 3 protons spin the complex once, which builds 1 ATP

ATP Synthase Structure

F0 channel is the rotor, gamma sticks into F1, transmembrane, (12 proteins), F1 is the stator, inside mitochondria, where the ATP is formed (in B subunit, which there are 3 of)

Stages of ATP Synthesis

Three sites as there are three beta proteins: one has ADP+Pi, one has formed ATP, one has released ATP. 1 full rotation is 3 H+.

Alternate uses of H+ gradient

To get ATP out of mitochondria, to get Pi and ADP into mitochondria

Starvation

Begins once all food is digested, body is now reliant on blood/storage, so activate glycogenolysis, lipolysis, and eventually proteolysis

Euglycemia

Need to have blood glucose above 4mM (ideally 5 mM)

Normal brain energy usage

120 g glucose/day

Body glucose requirements

Kidney, skin and RBCs have obligatory usage of glucose, as well as brain

Glycogenolysis

Glycogen mobilisation, phosphorylase breaks off a piece of glucose 1-phosphate (at the nonreducing-end), gets converted into G6P, G6Pase converts G6P into glucose, which enters blood via GLUT-2

Glycogen

Very branched, centre is glycogenin, each chain has 12-14 glucose residues, allows for quicker glycogenolysis

Glycogen phosphorylase

Breaks down glycogen, active when phosphorylated, active by phosphorylase kinase, deactivated by PPI

Glucagon Activity on Phosphorylase

Glucagon binds, triggers adenylyl cyclase, cAMP cascade activates PKA, which activates phosphorylase kinase, which activates glycogen phosphorylase

Phosphorylation Cascade

One small start e.g. glucagon can result in a massive amplification via phosphorylation of enzymes (typically activates them)

Debranching Enzyme

Transfers small branches to ends of other branches, allows phosphorylase to keep working, and also removes final branch component as glucose

Muscle Blood Glucose

Don't have G6Pase, so can't make normal glucose, stays in muscle cell

WAT Lipolysis

Hormone sensitive lipase and perilipin can break triacylglycerol into fatty acids, to be released into the blood

Hormone sensitive lipase

Activated by PKA, converts triacylglycerol into fatty acids and glycerol

Perilipin

Activated by PKA, encapsulates lipid droplet, when activated allows HSL to break down the fat

Glucagon Activity on Lipolysis

Causes phosphorylation cascade, cAMP, PKA, which activates perilipin and HSL

Starvation impact of FA Oxidation

Creates high levels of acetyl CoA, which increases PDH kinase activity, which deactivates the PDH, allowing glucose to stay at lactate and return to liver for gluconeogenesis

PDH Activation

High levels of Acetyl-CoA activate PDH kinase, which deactivates PDH, and insulin activates PDH phosphatase, which activates PDH