Teaching the design principles of metabolism

Abstract

Learning metabolism inevitably involves memorizing pathways. The teacher’s challenge is to motivate memorization and to help students progress beyond it. To this end, students should be taught a few fundamental chemical reaction mechanisms and how these are repeatedly used to achieve pathway goals. Pathway knowledge should then be reinforced through quantitative problems that emphasize the relevance of metabolism to bioengineering and medicine.

Figure 1

Reactivity of bonds β to carbonyl. (a) Protons β to carbonyl are acidic because the conjugate base is resonance stabilized. The same resonance stabilization also renders C-C bonds β to carbonyl labile. B, generic base. (b) Protons α to carbonyl cannot be abstracted because the resulting conjugate base has no stable resonance forms. By the same logic, C-C bonds α to carbonyl cannot be broken or formed without cofactor involvement. (c) For glucose, C-C bond cleavage β to carbonyl would result in products with unequal numbers of carbon atoms, whereas for fructose it yields two trioses. In glycolysis, glucose is isomerized to fructose before it is cleaved. (d) Isomerization of citrate to isocitrate in the TCA cycle positions a hydroxyl group such that its subsequent oxidation to carbonyl enables production of α-ketoglutarate by C-C bond breakage β to carbonyl. (e) Decarboxylation of pyruvate requires C-C bond breakage α to carbonyl. This is enabled by addition of the cofactor thiamine to pyruvate’s carbonyl, which repositions the scissile C-C bond β to the ‘pseudocarbonyl’ (blue) of thiamine’s thiazolium ring.

Figure 2

Example networks for teaching flux balance analysis. (a) Toy network. Students should learn to write down the flux balance equations shown to the right. They should then figure out, given F1 × 10 mmol h⁻¹ and that all fluxes are positive (or zero), what is the maximum possible rate of synthesis of the tetramer DCDE (F_objective). Note that the efflux from D to polymer synthesis is 2 × F_objective because each tetramer contains two D monomers. The answer is a discrete value (2.5 mmol h⁻¹) and can be achieved via multiple different sets of internal fluxes, always with F2 = 10 mmol h⁻¹ and F8 = 0. (b) Metabolic network of biofuel relevance. Students should figure out the values of X and Y (7 and 14, respectively), write down the differential equations for all metabolites and set them equal to zero, and then maximize F_objective. In this example, NADPH and ATP are assumed to be the sole cofactors for simplicity. AcCoA, acetyl CoA; FA, fatty acid.

Figure 3

The cancer-associated pyruvate kinase isozyme (PKM2) offers a modern case study for teaching metabolic regulation by isozyme switching, allostery and covalent modification. PKM2 turns off in response to tyrosine kinase signaling, oxidative stress and low serine concentrations, thereby promoting flux through the oxidative pentose phosphate and serine biosynthetic pathways (bold arrows). These fluxes, in turn, drive tumor growth. Solid lines indicate metabolic fluxes (for simplicity, stoichiometry and some cofactors and reactions are not shown). Dashed lines indicate regulation. The mechanism by which pyruvate kinase inhibition increases pentose phosphate pathway flux remains unknown. Challenging students with unanswered questions can motivate them to engage in metabolism research. G6P, glucose 6-phosphate; F6P, fructose 6-phosphate; FBP, fructose 1,6-bisphosphate; GAP, glyceraldehyde 3-phosphate; 3PG, 3-phosphoglycerate; PEP, phosphoenolpyruvate; ribose-P, ribose 5-phosphate.