Chapter 10: Introduction to Metabolism Metabolites: small molecules that are intermediates in the degradation or synthesis of biopolymers. Anabolic reactions: Synthesis of molecules necessary for the life of the cell. Catabolic reactions: Degradation of molecules to smaller molecules and to produce energy. The entire network of chemical reactions carried out by the cell AA, carb, nucleotides, fatty acids Animals need food for organic mol Provided by biosyn pathway in another species 4 major groups of biomolecules:

Metabolic Pathways Pathways are sequences of reactions: Linear pathway Cyclic pathway (citric acid cycle) Spiral pathway (fatty acid synthesis) Series of independent enzymesIntermediates are regenerated every turn (Very few are cyclic pathways) Same set of enzymes are used repeatedly Product is sub for next rxn Polymerization rxns

Major Metabolic Pathways Catabolic Pathways Reactions and pathways can be linked to form extended metabolic routes

Major Metabolic Pathways Autotrophs: can utilize inorganic sources of essential elements. Anabolic Pathways (biosynthesis) Photoautotrophs Chemoautotrophs Large mol from smaller mol By adding C and N Energy provided by Light or Breakdown of organic mol from other autotrophs Heterotrophs: need mol such as glucose Most biochemically complex organisms?? Autotrophs

Catabolic Pathways Not simply reverse of anabolic rxns AA, nucleotides, monosacc, FA are formed by hydrolysis Then degraded in oxidative rxns and energy conserved in ATP and red coenzymes (NADH) Eliminate unwanted mol and generate energy CAC: main source of energy to drive ATP syn But also important in anabolic pathways: source of precurcors for AA syn Main role:

Metabolism Proceeds in Discrete Steps Narrow conditions of the cell (pH, temperature, pressure, concentrations) require specific and efficient enzymes. Limited reaction specificity of enzymes requires multiple steps. Multiple steps required to control energy input and output. Allow sharing of intermediates. Allow more control points to regulate biochem process. But why are there so many distinct rxns involved ?? Energy carriers (ATP/NADH) are in all life forms

Metabolic Pathways are Regulated Organisms react to changing environmental conditions (avail. of nutrients) and genetically programmed instructions (during development) Regulate synthesis/degradation of biomol and generation/ use of energy Most pathways: Go in single direction (in physiological conditions) Without backing up and wasting energy etc Flux = flow through a pathway. Flow will continue as long as A is high and P is removed Intermediates are in steady state (B,C,D,E do not change much) Special regulatory controls that affect particular enzymes in pathway Usually there are multiple control points “regulatory enzymes”

Regulation of Metabolic Pathways Feedback inhibition. Feed-forward activation. Controls the first committed step (the first rxn unique to pathway) Prevent unnecessary accumulation of metabolic intermediates Inc in metabolite B increases flux thru the pathway Two patterns of metabolic regulation Allosteric modulation

Regulation of Glycolysis Signals adequate supply of energy Intermediate of CAC Glycolysis activity dec when its products are no longer reguired First committed step of glycolysis High AMP indicates dec. ATP

Phosphofructokinase-1 ATP: allosteric inhibitor AMP: allosteric activator It is an allosteric enzyme and a regulatory step for glycolysis Metabolically irreversible rxn. [ATP] > ADP / AMP

Regulation of Metabolic Pathways Regulation of pathway occurs through allosteric modulators (inhibitors and activators), and by covalent modification (protein kinases,phosphatases). Amplification of original signal Kinases with multiple specificities: Coordinated reg of many pathyways Slow process : relative to allosteric/covalent Anabolic pathways: phosp-inactive Rapid and reversible Catabolic pathways: phosp-active Regulated rate of enzyme syn/ degradation phosphorylated protein is active or inactive

A + B C + D K eq = [C][D] [A][B]  G = 0 at equilibrium, no net synthesis of products  G reaction =  G o’ + RTln [C][D] [A][B] When  G reaction > 0, the reaction is unfavorable, energy needed to make  G neg.. When  G reaction << 0, the reaction is favorable, spontaneous and irreversible (no external source of energy needed). The Free Energy of Metabolic Reactions Gibbs free energy change: measure of energy available from a rxn Standard Gibbs free energy: change under standard conditions (1M reactants and products) Actual Gibbs free energy: depends on real concentrations  G =  H - T  S chem process) Measure how far from equilibrium the reacting system is operating  G o’ GG Determine the spontaneity of a rxn and thus its direction GG

A + B C + D K eq = [C][D] [A][B]  G = 0 at equilibrium, no net synthesis of products  G reaction =  G o’ + RTln [C][D] [A][B] Gibbs free energy change Measure how far from equilibrium the reacting system is operating Near equilibrium rxns: small free energy changes Rxns are readily reversible Can accomodate flux in either direction and quickly restore levels of R and P to equil status (Most metabolic rxns) Not suitable control points of a pathway Direction of rxn can be controlled by changes in [S] [P] GG

A + B C + D K eq = [C][D] [A][B]  G = 0 at equilibrium, no net synthesis of products  G reaction =  G o’ + RTln [C][D] [A][B] Near equilibrium rxns: small free energy changes Metabolically irreversible rxns Rxns are readily reversible Rxns greatly displaced from equil Large  G Can accomodate flux in either direction and quickly restore levels of R and P to equil status Flux is is unaffected by changes in metabolite conc thus usually controlled by modulating the enzyme (Most metabolic rxns) Gibbs free energy change Measure how far from equilibrium the reacting system is operating GG

Metabolic Pathways are Regulated Most pathways: Go in single direction Without backing up and wasting energy etc Regulation of unidirectional rxns (irreversible) “regulatory enzymes” [S] and [P] far from equilibrium Regulatory enzyme controls flux E6E6 Reverse reaction need different enzyme…….key regulatory step unaffected by changes in metabolite conc controlled by modulating the enzyme Large  G

Metabolically irreversible rxns : need diff enzyme for reverse Fructose 1,6 bisphosphatase Inhibited by AMP Inc in AMP indicates Dec in ATP Need ATP Need glycolysis Inhibit glc synthesis

The Free Energy of ATP ATP (and PPi) is an energy- rich compound: Why large amt of energy released during hydrolysis? Electrostatic repulsion of negative charges on phosphoanhydride bonds is less after hydrolysis. Products of hydrolysis are better solvated (by H 2 O) than ATP. The products of hydrolysis are more stable than ATP. e- on terminal oxygens more delocalized Phosphoanhydride vs Phosphoester linkage Dec the repulsion of P groups drive the rxn Bridging O two terminal O

The Free Energy of ATP [ATP] >> other NTP but all are called ‘energy rich compounds’ Intracellular [ATP] little fluctuation Maintained by adenylate kinase AMP + ATP + 2Pi 2 ATP + 2 H20 [ATP] > ADP / AMP small change in [ATP]…large change in [ADP/AMP] allosteric modulator of some energy yielding processes Phosphoanhydride higher energy

The Metabolic Roles of ATP Phosphoryl-group transfer: Glutamate + NH 4 + Glutamine;  G o’ = +14 kJ mol -1 Rxn not possible (not spontaneous) in living cells: Glutamine steady state levels are high Limiting supply of ammonia ATP hydrolysis drives the rxn

The Metabolic Roles of ATP Phosphoryl-group transfer: Glutamate + NH 4 + Glutamine;  G o’ = +14 kJ mol -1 Glutamate + ATP Glutamyl-P + ADP Glutamyl-P + NH 4 + Glutamine +P i Glutamate + NH 4 + Glutamine  G o’ = +14 kJ mol -1 ATP ADP + P i  G o’ = -32 kJ mol -1 Glutamate + NH ATP Glutamine + ADP + Pi  G o’ = -18 kJ mol -1 Carboxyl group Is activated Coupled rxn (Glutamine synthetase)

Phosphoryl group-transfer potential ATP: intermediate phos-group-trans potential and kinetically stable thus mediates most energy transfers Measure of free energy required for formation of the phosp compound

Production of ATP by phosphoryl group transfer: Phosphagens High phosphoryl group-transfer potential More stableMetabolically irreversible Energy storage in muscle 3-4 sec burst of energy When ATP low Allows replenishment

The Metabolic Roles of ATP Nucleotidyl group transfer: ATP AMP + PP i Acetate + HS-CoA Acetyl-CoA PP i 2P i  G o’ = -45 kJ mol -1  G o’ = +32 kJ mol -1  G o’ = -29 kJ mol -1 Acetate + ATP + HS-CoA Acetyl-CoA + AMP + 2P i  G o’ = -42 kJ mol -1 Removal of (PPi) product drives the rxn AMP is transferred

Thioesters have high free energies of hydrolysis GTP synthesis by coupling to thioester hydrolysis. Thioesters less stable than oxygen esters unshared e- of S not effectively delocalized Energy similar to phosphoanhydride linkage Substrate level phosphorylation Conserves the energy used in formation