Quiz #4/5

#4: Glycolysis (Tuesday, Feb 20 th ) #5: TCA cycle (Monday, Mar 5 th ) Pathways are in the books Quiz will have the entire pathway: –All cofactors will be present –Random intermediate and enzymes removed You fill in the missing names –Draw the structure for 1 intermediate Indicated by a larger box

Enzyme Regulation

Conditions Affecting Enzyme Activity pH temperature

pH

Effects of pH on Enzyme Activity Protonation state of side chains –Variation in protein structure –Substrate binding –catalysis Ionization of substrate –Substrate binding

Temperature Protein unfolding

Control of Enzyme Availability Principles of Genetic Regulation

Types of Enzymes “ Control of Gene Expression” Constitutive Enzymes: e.g. glycolytic enzymes and gluconeogenic enzymes Inducible Enzymes: e.g.  -galactosidase Repressible Enzymes: e.g. ten enzymes of histidine biosynthesis

Negative Regulators [Bind to operators or upstream repression sequences (URS)]

Positive Regulators [Bind to promoters, enhancers or upstream activation sequences (UAS)]

Regulation of Enzyme Catalytic Activity Covalent Modification Allosteric Enzymes

Principles Governing Controls of Enzyme Catalytic Activity Regulatory Enzymes –Enzyme catalyzing committed, rate-limiting step (often first step) –Thermodynamically highly favorable reaction Outcomes of Regulation –Feedback inhibition (fbi) of biosynthetic pathways –Modulation of metabolic flux

Reversible Covalent Modification

Page 390 Protein Modification (Phosphorylation/Dephosphorylation)

Non-covalent Modification Effectors or Ligands Positive: activators Negative: inhibitors

Allosteric Enzymes (Modulation of Enzyme Catalytic Activity) Substrate Binding Catalytic Rate Both

Allosteric (Regulatory) Enzymes

Homotropic Effects

Heterotropic Effects

Figure Glycogen Phosphorylase

Regulation of Biosynthetic Pathways

Rationale for Regulation Efficiency and Flexibility

Biological Efficiency Biosynthesis –Synthesize precursors not available in diet –Cease synthesis when precursors become available in diet (pre-existing enzymes) –Produce precursors and macromolecules at appropriate rates Catabolism –Degrade most appropriate nutrients at appropriate rates

Biological Flexibility Adaptaton to Dietary Changes –Need for biosynthetic products –Catabolism of new nutrients –Control of pre-existing enzymes Metabolic Flux –Rates of metabolism reflecting needs for energy and macromolecular synthesis

Competing Reactions: Regulation

Control Mechanisms Control of Enzyme Availability –Induction/repression Control of Enzyme Activity –Covalent/Non-covalent Control of Substrate Availability

Types of Regulation Specific: pathway’s substrate or product General: needs for C or N sources or growth rates (e.g. energy charge)

Signals Mediating Regulation Availability of Substrates or Products (Ligands) Regulatory Proteins

Biosynthetic Pathways

Simple Feedback Inhibition

Complex Feedback Inhibition

Mechanisms of Complex Feedback Inhibition Cumulative: sum of individual inhibitions Concerted: both end products required for inhibition Isoenzyme: two enzymes, each inhibitable by different end product

Cumulative Feedback Inhibition A GF ED CB A GF ED CBA GF ED CB

Concerted Feedback Inhibition A GF ED CB A GF ED CBA GF ED CB

Isozymes A GF ED CBA GF ED CB A GF ED CB

Modulation of Metabolic Flux Energy Charge

Energy Charge (Daniel Atkinson) Steady-State E.C. = 0.93 ATP, ADP and AMP = Regulatory Ligands

Energy Charge Anabolic pathways (Biosynthesis) Require ATP Activated –High EC (ATP) Inhibited –Low EC (AMP) Catabolic Pathways (Degradation) Produce ATP Activated –Low EC (AMP) Inhibited –Hig EC (ATP)