1. Metabolism Collection of biochemical rxns within a cell Metabolic pathways –Sequence of rxns –Each step catalyzed by a different enzyme Enzymes of a pathway often physically interact to form large complexes –Limits amount of diffusion needed at each step of the pathway –The product of the preceding step is the reactant in the following step –Metabolic intermediates are the products formed along the way towards the ‘final’ product oxaloacetate

2. Catabolism vs Anabolism Catabolic: breakdown from complex to simple –Yield raw materials (amino acids, etc) and chemical energy (NADH, ATP) –Convergent: diverse starting materials broken down to conserved set of intermediates (pyruvate, Acetyl-CoA) Anabolic: synthesis from simple to complex –Consume raw materials and chemical energy stored in NADPH and ATP –Divergent: small set of molecules assembled into a diversity of products

3. Catabolism vs Anabolism Catabolism Anabolism

4. Oxidation and reduction Redox reactions: the gain (reduction) or loss (oxidation) of electrons –Reducing agents = lose e- = get oxidized –Oxidizing agents = gain e- = get reduced Fe 0 + Cu 2+ Fe 2+ + Cu 0 Reducing agent + oxidizing agent oxidized + reduced –Metals show complete transfer of e- Reducing agents reduce the charge on oxidizing agents

5. Oxidation and reduction Redox reactions: the gain (reduction) or loss (oxidation) of electrons –Changes in organic molecules shift the degree of e- sharing Carbon in C-H bond is reduced Carbon in C=O bond is oxidized –EN diffs result in e- spending less time around C when bonded to O CH4 + 2O2 --> CO2 + 2H2O

6. Capture and Use of E Alkanes are highly reduced organic compounds (E rich) –Not well tolerated by most cells Fatty acids and sugars are well tolerated C 6 H 12 O 6 + 6O 2 --> 6CO 2 + 6H 2 OΔG°’= -686 kcal/mol ADP + Pi --> ATPΔG°’= +7.3 kcal/mol Theoretical Yield ~ 93 ATP Actual (aerobic) ~ 36 ATP39% efficient –Marathon runner Actual (anaerobic)= 2 ATP2% efficient –Sprinter

7. Glycolysis Glucose + 2NAD + 2ADP + 2Pi --> 2pyruvate + 2ATP + 2NADH

8. K’eq ΔG°’ ΔG for actual cell conditions

9. Kinase: an enzyme that can transfer phosphate from ATP to another molecule Phosphatase: hydrolyzes phosphate from a molecule Isomerase: an enzyme that can catalyze structural rearrangements Steps 1-3: 2 ATP used

10. Aldolase: an enzyme that cleaves an aldol (which is a beta-hydroxy ketone)

11. Two modes of E extraction 1. Extraction of H+ and 2e- (:H - ) –NAD + + H: --> NADH –Extraction of :H - is done by dehydrogenase enzymes Dehydrogenase: oxidizes substrates by transferring hydride (H-) ions to an electron acceptor (e.g. NAD+).

12. Nicotinamide Adenine Dinucleotide (NAD) add :H - to the nicotinamide ring Most NADH destined for electron-transport chain Add phosphate to ribose 2’-OH creates NADP/NADPH rAMP

13. Another example of an ES complex with a covalent intermediate Regenerate enzyme in last step using inorganic phosphate (Pi)

14. Two modes of E extraction 2. Substrate level phosphorylation of ADP --> ATP –transfer of phosphate from higher energy compounds to lower energy ones ATP is not the highest energy compound Reverse reaction looks like a classic kinase

15. Mutase: shifts the position of a functional group aka as a hydratase

16. Glycolysis: summary Steps 1, 3 –2 ATP consumed Step 4 –6C sugar split into two 3C sugars Step 6 –Redox reaction: NAD + + :H - --> NADH Step 7, 10 –Substrate level phosphorylation Glucose + 2NAD + + 2ADP + 2Pi --> 2Pyruvate + 2ATP + 2NADH No O2 used, anaerobic

17. Reducing power: NADH vs NADPH Synthesis of fats from sugar requires reduction of metabolites H-C-OH + :H - + H + ---> H-C-H + H2O NADH is generated from Catabolic pathways NADH + NADP + NAD + + NADPH transhydrogenase NADPH is used as reducing agent for Anabolic pathways

18. Fermentation can regenerate NAD + Under anaerobic conditions –Skeletal muscle: Pyruvate + NADH ---> Lactate + NAD + –Yeast: Pyruvate ---> Acetaldehyde + CO2 Acetaldehyde + NADH ---> Ethanol + NAD + - O2

19. Fermentation can regenerate NAD + Under anaerobic conditions –Skeletal muscle: Pyruvate + NADH ---> Lactate + NAD + –Yeast: Pyruvate ---> Acetaldehyde + CO2 Acetaldehyde + NADH ---> Ethanol + NAD + Under aerobic conditions –Pyruvate enters TCA cycle –NAD+ regenerated by electron transport chain (oxidative phosphorylation) + O2

20. Regulation of enzyme activity Allosteric modulation (Allostery) –Binding of a molecule to the enzyme activates or inhibits it –Binding occurs at an ‘allosteric site’ on the enzyme –Feedback inhibition: Final product of a pathway inhibits the first enzyme in the pathway Keeps level of product from getting higher than needed A + B --> C + D --> E E is an allosteric inhibitor that binds to allosteric site blocking 1st rxn

21. Allosteric regulation of metabolism Most cells have enzymes for both glycolysis and gluconeogenesis Allostery controls which pathway is active versus inhibited to provide sensitivity to energy needs

22. Allosteric regulation of metabolism AMP = allosteric inhibitorATP = allosteric inhibitor AMP = allosteric activator ATP --> ADP + Pi ADP + ADP --> ATP + AMP

23. Regulation of enzyme activity by covalent modification Phosphorylation uncharged charged –SerineH 2 C-OH --> H 2 C-O-PO 3 2- –Threonine also subject to phosphorylation –Tyrosine also subject to phosphorylation –These subtle changes to the chemical information guiding protein folding can yield conformational changes in protein structure that increase or decrease enzyme activity enz P protein phosphatases protein kinases

24. Metabolism: cell overview