biochemistry S2 Y1 Flashcards
Role of non-photosynthetic energy conversion pathways?
Catabolise carbon-based fuels (fats, carbs and proteins) to reduce O2
What does the inner mitochondrial membrane contain to enable the ETC to occur?
Complexes for electrons to pass through
Why is there a proton gradient across the inner-mitochondrial membrane?
Electrochemical movement of protons from area of high conc. to low conc. to produce ATP (due to redox running electron motive force)
2 things the proton gradient establishes?
- pH gradient that forms chemical potential energy
- Electrical difference that forms electrical potential energy
Why is low pH created in mitochondrial matrix?
Protons then move into the matrix through ATP synthase to form ATP (proton flow = electric current, ATP synthase = resistor)
Why is low pH created in chloroplasts’ cytosol?
Photosystems move protons into thylakoids and then they move out to produce ATP
What is H+ uncoupling?
Generation of heat rather than ATP
What are blocking effects?
Blockers shutting down H+ flow to cause cell death as DPH and DY increase
When is proton uncoupling used?
In hibernation, newborn mammals, cold-adapted animals and bodybuilders to generate heat
Why have humans used 2,4-DNP?
It is an uncoupler that causes rapid weight loss
How was the chemiosmotic theory proven?
Using an artificial membrane containing a bacteriorhodopsin protein (much like a photosystem - light lets H+ through) and a mitochondria - ATP WAS SYNTHESISED
What carry out oxidative phosphorylation?
Four protein complexes (I-IV), cytochrome C, ATP synthase
What provide channels for small molecules across outer membranes?
Porin proteins
Role of translocase proteins?
Shuttle ATP, ADP, Pi across inner-mitochondrial membrane
- Number of H+ translocated at the start of the ETC?
- How many reenter per ATP?
- 10
- 4 (3 via ATP synthase, 1 via phosphate translocase)
What happens to the electron donor whilst transferring to acceptor (the oxidant)?
It is oxidised (acts as a reductant)
Do oxidation and reduction occur simultaneously or at different times?
Simultaneously (as shown by half-equations, oxidised form on LHS)
4 ways electrons are transferred from donor to acceptor?
- Directly as electrons
- As hydrogen atoms
- As a hybride ion (:H-)
- Direct combination with oxygen
- What depends on redox potential?
- How is standard redox potential of a couple measured?
- Role of strong reducing agent (e.g. NADH)?
- Role of strong oxidising agent (e.g. Fe3+)?
- Tendency of a redox couple accepting and donating
- Using electrochemical cell relative to standard hydrogen electrode
- Poised to donate electrons (has negative redox potential)
- Ready to accept electrons (has positive redox potential)
What is E°’?
Potential of a redox couple in which reduced and oxidised species are present at 1M, 25°C, pH7
- Where do electrons flow in a spontaneous reaction?
- ΔG°’ and ΔE°’ in spontaneous reaction?
- From redox couple of lower potential to redox couple of higher potential
- ΔG°’ is negative
ΔE°’ is positive
Equation for work done when an electron is moved in an electric field?
Work done = electron charge x potential
Equation for ΔG°’?
ΔG°’ = -n x F x ΔE°’
n = no. of electrons transferred
F = Faraday constant (96.5)
ΔE°’ = difference in standard reduction potentials between 2 redox couples (V)
What is the Nernst equation?
E’ = ΔE°’ + (2.303 RT / nF) log10 ([e- acceptor] / [e- donor])
- At 25°C, (2.303 RT / nF) = 0.059 for 1 electron transfer, 0.0295 for 2
Mitochondrial electron transport system:
- What is it?
- Where do electrons from NADH flow?
- How many H+ translocated concomitantly?
- A series of coupled redox reactions
- Enter complex I, then CoQ, complex III and complex IV
- 10
Mitochondrial electron transport system:
- Where electron pairs derived from FADH2?
- How many H+ translocated concomitantly?
- Complex II, ETF-Q oxidoreductase or glycerol-3-phosphate dehydrogenase
- 6
Complex I (NADH-ubiquinone oxidoreductase):
- What is coupled?
- How many Fe-S carrying e- at a time?
- What can Fe-S clusters change?
- How many H+ translocated into intermembrane space?
- NADH oxidation (releases 2e-) and FMN reduction (forms semiquinone intermediate if it is 1e- and FMNH2 if it is 2e-)
- 7
- Redox potential from -0.5 to 0.4V (depending on protein microenvironment)
- 4
Coenzyme Q (ubiquinone):
- 3 roles?
- Mobile, lipid-soluble e- carrier that transports electrons in membrane from complex I to III
- Entry point into ETC for e- pairs from CAC, fatty acid oxidation and glycerol-3-phosphate dehydrogenase
- Converts 2e- transport system in complexes I and II to 1e- system in complex III (which then passes electrons to cytochrome C one at a time)
Complex II (succinate dehydrogenase):
- What is it linked to?
- Role?
- CAC
- Oxidises succinate to fumarate (coupled to FAD/FADH2) – electron pair then used to reduce Q to QH2 via Fe-S and a haem – electrons move through Fe-S clusters and cytochrome b560 (FADH2 then oxidised)
Complex III (ubiquinone-cytochrome c oxidoreductase):
- What 2 binding sites?
- What are its prosthetic groups for?
- Role?
- What kind of complex?
- Why does CoQ utilise the Q cycle?
- Q binding sites (Qp and QN)
- Function as electron carriers
- Reduces cytochrome c and translocates 4H+
- Dimeric (2x11 subunits)
- Converts 2e- process into 2 x 1e- transfers
Haems and cytochromes:
- How is cytochrome c haem group linked to protein?
- What does a type a haem have?
- Covalently through thiol groups from cysteine residues
- Long hydrophobic tail
Complex IV (cytochrom c oxidase):
- Role?
- What is electron transport though?
- What happens to 4 H+?
- Cytochrome c oxidation
- Through one monomer of homodimer (culminating O2 reduction to form H2O)
- 2H+ translocated into inter-membrane space, 2H+ used top form H2O
What do complexes I-IV all have?
Transmembrane regions and functional domains protruding into the matrix (III and IV have functional domains that protrude into the intermembrane space to interact with cyt c)
How many H+ does NADH oxidation starting at complex I translocate?
10
How many H+ does CoQ + FADH2 oxidation starting at complex II translocate?
6
Why must CoQ and cyt c make 2 trips to transfer 2e- from NADH and FADH2?
Can only transfer 1e- at a time
Reactions for reactive oxygen species and H+ making water?
O2-. + 2H+ <–> O2 + H2O2 (SOD catalyses)
2 H2O2 <–> O2 + H2O (catalase catalyses)
Equation for the transfer of 2e- from NADH through respiratory chain to O2?
ΔG°’ = -nFΔE°’ = -nF[E°’(acceptor) - E°’(donor)]
Equation for ΔG available from H+ gradient across mitochondrial membrane?
ΔG = RT ln (c2/c1) + ZFΔΦ
- Z is absolute value of its charge
- F is faraday constant
- Φ is electrical potential difference across
membrane
ΔpH and ΔΦ in actively respiring mitochondria?
ΔpH = 0.75
ΔΦ = -0.15V
Is more free energy available from H+ gradient derived from ΔpH or ΔΦ?
ΔΦ
What subunits do all ATP synthases have?
3 alpha and 3 beta
What is different about bacterial ATP synthase?
Has gamma subunit that associates with F1 component as the 3 alpha and beta subunits
What is different about yeast ATP synthase?
Has the gamma subunit, 3 alpha and beta, a delta subunit homologous to bacterial gamma subunit and an OSCP subunit
What are the F0 and F1 components for in ATP synthase?
F0 is a H+ channel, F1 is for catalytic activity
What is the stator of ATP synthase?
Subunit with half-channels for H+ to enter and exit AND a stabilising arm (b, d, h + OSCP)
What is the rotor of ATP synthase?
c + g + d + e rotate as H+ enter and exit the c-ring
What is the headpiece of ATP synthase?
Hexameric a3b3 unit responsible for ATP synthesis
What are the 3 basic principles of ATP synthase?
- Gamma directly contacts all 3 beta subunits, but each interaction is distinct (gives rise to 3 different beta conformations)
- ATP binding affinities of the 3 beta subunits are T, L, O (tight = ATP bound, loose = ADP + Pi bound, open = ATP released)
- H+ flow through F0 cause rotation of gamma subunit counter-clockwise during ATP synthesis - each 120° rotation cuases beta subunits to go fro, L –> T –> O –> L etc
What direction does ATP synthase rotate in?
In reverse conditions that favour ATP hydrolysis
How can ATP synthesis be catalysed?
a3b3 headpiece simply as the function of the g subunit is imposed by electromagnets
What are disaccharides attached by?
O-glycosidic bonds
What do alpha and beta glucose react to form?
Maltose
2 examples of reducing sugars?
Maltose and lactose
Example of non-reducing sugar?
Sucrose
What are the two types of oligosaccharide?
- 2-8 linked monosaccharides (disaccharides included)
- 3-8 saccharide molecules (low abundance)
What are polysaccharides?
> 8 saccharides for structure or storage
2 examples of structural polysaccharides?
- Cellulose (beta (1-4) linked glucose units)
- Chitin (beta (1-4) linked N-acetylglucosamine units that are linked and unbranched)
What does beta 1-4 linkage of glucose form?
Straight chains whereby hydrogen bonds can form with adjacent molecules to add further stability
2 examples of polysaccharides for storage?
- Starch - alpha amylose (unbranched alpha 1-4 glucose polymer) and amylopectin (branched and linked alpha 1-6 glucose polymer)
- Glycogen (similar to amylopectin but branches 3x more)
What do alpha 1-4 linkages cause?
Coiled chains (twist around one another)
Glycosaminoglycans (GAGs):
- What are they?
- Major role?
- Example?
- Polymers composed of repeating disaccharide units
- Component of ground substance (holds tissue types together)
- Hyaluronic acid
Hyaluronic acid:
- Where?
- Made up of?
- How are disaccharides attached?
- Characteristics?
- Ground substance, synovial fluid, vitreous humour of eyes
- D-glucuronic acid and N-acetyl-D-glucosamine
- Beta 1-4
- Rigid, highly hydrated, viscous, absorbs shock and shearing forces
What are glycoproteins?
Proteins modified through the addition of carbohydrates without strict genetic control
What determines the carbohydrate added to proteins?
Enzymes available at the time = microheterogenity
2 types of carbohydrate attachment to proteins in glycoprotein formation?
- N-linked
- Needs amino acid side chain with N
- E.g. N-linked-N-acetylglucosamine is beta
linked to nitrogen of Asn - O-linked
- OH group of Ser or Thr (sometimes
HyrdroxyLys) attachment
- More variable than N-linked
What is glycogen composed of?
Alpha 1-4 linked chains and alpha 1-6 linked branches
What does the glycogen chain start with?
Glycogenin proteins (where A and B chains begin)
THERE ARE TWO IDENTICAL PROTEINS
Act as primer for glycogen formation
What are the inner and outer regions of glycogen?
Inner = B-chains with 2 branch points
Outer = A-chains that aren’t branched
How does glycogen branch?
When 13 residues have been added to growing strand, branching enzyme recognises this and make a branch to grow a new chain
Why are the outermost chains of glycogen unbranched?
Makes glucose more easily accessible (outermost tier always contains 34.6% of glucose in structure)
What are other proteins associated with glycogen for?
Synthesis and breakdown
How does glycogenin act as a catalyser of glucose addition?
First glucose added to Tyr195 and then subsequent glucoses are added to growing chain (until 10-20 residues when glycogen synthase takes over)
What do glycogenin and glycogen synthase utilise?
Activated precursors (UDP-glucose in eukaryotes, ADP-glucose in bacteria and plants)
UTP + glucose-1-phosphate –> UDP-glucose
CATALYSED by UDP-glucose pyrophosphorylase
Role of UDP-glucose pyrophosphorylase?
Cleaves alpha and beta phosphate in UTP
Role of inorganic pyrophosphatase?
Hydrolyses the phosphates released from UTP in the cytoplasm to prevent the reverse reaction
What type of reaction is UDP-glucose addition?
Glycosyl transfer with the release of UDP via double nucleophilic substitution or intramolecular nucleophilic substitution
What is the glycogen branching enzyme and what does it do?
Amylo-(1,4 –> 1,6) transglycosylase
Transfers terminal chain section (around 7 residues) to the C6-OH of another glycogen chain
What enzymes degrade glycogen?
Glycogen phosphorylase
Glycogen debranching enzyme Phosphoglucomutase
- What does glycogen phosphorylase do?
- Mechanism?
- Hydrolyses 1-4 bond which releases alpha-D-glucose-1-phosphate and glycogen
- Via a carbocation (Sn1) –> carbocation stabilised by pyridoxal phosphate which is covalently linked to the enzyme (PLP is active for of vitamin B6)
Debranching enzyme:
- 2 functions? (bifunctional)
- Mechanism in notes, but what is the result?
- Transferase and alpha-1,6-glucosidase
- No branch
Phosphoglucomutase:
- Role?
- Fates of glucose-6-phosphate?
- What happens in periods of plentiful glucose?
- Converts glucose-1-phosphate to glucose-6-phosphate
- Enters glycolysis OR dephosphorylated to form glucose in the liver
- G-6-P formed by hexokinase to change the equilibrium position so that the enzyme converts G-6-P into G-1-P that can form UDP-glucose for glycogen synthesis
How does the relationship between the synthetic capacity and the degrative capacity vary between the liver and muscle?
Liver = roughly equal
Muscle = degradation can be 300x faster
How is glycogen metabolism regulated?
Through synthesis (using glycogen synthase) and degradation (using glycogen phosphorylase) –> regulated by phosphorylation and dephosphorylation
Signalling cascade of glycogen metabolism?
1 signal molecule –> 10 second messengers –> 100 molecules activated –> 1000 glucose released
How does insulin lower blood glucose?
Increases glycogen synthase activity (increases conversion from phosphorylated b form to dephosphorylated ACTIVE a form)
It does this by binding to an insulin receptor, phosphorylating insulin receptor substrate which interacts with PI3K to for PIP3 that signals the target GSK3 molecule and phosphorylates ut to deactivate it so glycogen synthase is activated
Glycogen synthase kinase 3 (GSK3):
- How does it inactivate glycogen synthase?
- When can the P be added?
- Phosphorylates serine side chains (has site where it recognises primed phosphate and a site where it adds the phosphate)
- After priming reaction that occurs via casein kinase II action
What catalyses dephosphorylation of glycogen synthase?
Phosphoprotein phosphatase 1 (PP1)
- acts quicker if G-6-P is bound to glycogen
synthase
What does PP1 activate and what does it inactivate?
Activates glycogen synthase
Inactivates glycogen phosphorylase and phosphorylase kinase
How is glycogen phosphorylase regulated?
As it is more active when it is phosphorylated, it is phosphorylated by phosphorylase kinase and dephosphorylated by phosphorylase phosphatase 1
2 ways glycogen is signalled to be degraded?
- Glucagon signals liver to release glucose into blood
- Adrenaline signals muscles to release glucose for energy production
5 steps of mechanism for signal for glycogen degradation?
- Either molecule signals GDP to GTP conversion
- GTP interacts with adenylate cyclase
- ATP converted to cAMP
- cAMP causes protein kinase A to activate phosphorylase kinase
- Glycogen phosphorylase is activated
2 way muscle glycogen phosphorylase is activated?
- Ca2+ released into muscles to induce contraction which stimulates phosphorylase kinase
- AMP builds up if not enough ATP is produced
What inhibits liver glycogen phosphorylase?
Glucose - glycogen phosphorylase acts as a glucose sensor - bound phosphates more exposed to removal by PP1
PP1:
- What does it increase?
- What does it decrease?
- What is it inhibited by?
- What is it bound to?
- Attached to?
- Glycogen synthase activity by dephosphorylation
- Glycogen phosphorylase by dephosphorylation
- PKA
- Glycogen with GS, PK and GP
- Glycogen-targetting protein
How are carbohydrate and lipid metabolisms regulated?
Co-ordinately
- Signalling factors have co-ordinated effects on glycogen synthesis and breakdown and glycolysis AS WELL AS lipid synthesis and breakdown
Why are simple lipids less common than complex ones?
Simple have more specific functions (e.g. cholesterol) and complex ones have greater range (e.g. phospholipids)
What are complex lipids made up of?
Smaller components - fatty acid is major component
What are fatty acids composed of?
Saturated/unsaturated long chain hydrocarbon (most double bonds are cis), and a carboxyl group (pKa = 4.5) and is ionised at most physiological pHs
What is the nomenclature of fatty acids?
Number of carbons in chain + ‘oic acid’ or ‘oate’ if ionised
Are all the R groups in triacylglycerols the same?
No, the differences determine the type of fat or oil
How does the structure of phospholipids differ from triacylglycerols?
Has a polar (hydrophilic) head group
How does chain length affect inter-chain interactions?
Longer chains create more interactions
How do double bonds affect inter-chain interactions?
More double bonds = less interactions
What does the extent of inter-chain interactions affect?
Fluidity levels
What are amphipathic molecules also known as?
Schizophrenic
Examples of amphipathic molecules?
Free fatty acids, phospholipids, glycolipids, sphingolipids and ceramides
- What do amphipathic molecules naturally form?
- What does vigorous shaking cause?
- Monolayers and sometimes bilayers
- Micelle formation (driven by entropy)
3 sources of fatty acids?
- Diet
- Adipose storage
- De novo synthesis
Where does fat digestion begin?
In the small intestine and uses the pancreas+liver
What produces and releases bile and what is its role?
Liver produces, gall bladder releases
As it is made from bile acids and salts derived from cholesterol, it acts as a dtergent (emulsifies lipids to form fat droplets)
What does the pancreas produce?
Digestive enzymes and bicarbonate solution
What enzyme does the pancreas release for fat digestion and how does it work?
Lipase
Breakdown micelles into monoglycerides and fatty acids, these are then incorporated into the ER as they move down the concentration gradient from the gut into cells –> triglycerides then synthesised, chylomicrons formed –> enter blood via lymphatic vessel