Harpers

Enzymes: Kinetics 8Victor W. Rodwell, PhD, & Peter J. Kennelly, PhDBIOMEDICAL IMPORTANCEEnzyme kinetics is the field of biochemistry concernedwith the quantitative measurement of the rates of enzyme-catalyzedreactions and the systematic study of factorsthat affect these rates. Kinetic analyses permit scientiststo reconstruct the number and order of theindividual steps by which enzymes transform substratesinto products. The study of enzyme kinetics also representsthe principal way to identify potential therapeuticagents that selectively enhance or inhibit the rates of specificenzyme-catalyzed processes. Together with sitedirectedmutagenesis and other techniques that probeprotein structure, kinetic analysis can also reveal detailsof the catalytic mechanism. A complete, balanced set ofenzyme activities is of fundamental importance for maintaininghomeostasis. An understanding of enzyme kineticsthus is important for understanding how physiologicstresses such as anoxia, metabolic acidosis or alkalosis,toxins, and pharmacologic agents affect that balance.CHEMICAL REACTIONS ARE DESCRIBEDUSING BALANCED EQUATIONSA balanced chemical equation lists the initial chemicalspecies (substrates) present and the new chemicalspecies (products) formed for a particular chemical reaction,all in their correct proportions or stoichiometry.For example, balanced equation (1) below describesthe reaction of one molecule each of substrates A and Bto form one molecule each of products P and Q.A+ B ← → P+QThe double arrows indicate reversibility, an intrinsicproperty of all chemical reactions. Thus, for reaction(1), if A and B can form P and Q, then P and Q canalso form A and B. Designation of a particular reactantas a “substrate” or “product” is therefore somewhat arbitrarysince the products for a reaction written in onedirection are the substrates for the reverse reaction. Theterm “products” is, however, often used to designatethe reactants whose formation is thermodynamically favored.Reactions for which thermodynamic factorsstrongly favor formation of the products to which thearrow points often are represented with a single arrowas if they were “irreversible”:(1)60A+ B → P+QUnidirectional arrows are also used to describe reactionsin living cells where the products of reaction (2)are immediately consumed by a subsequent enzymecatalyzedreaction. The rapid removal of product P orQ therefore precludes occurrence of the reverse reaction,rendering equation (2) functionally irreversibleunder physiologic conditions.CHANGES IN FREE ENERGY DETERMINETHE DIRECTION & EQUILIBRIUM STATEOF CHEMICAL REACTIONSThe Gibbs free energy change ∆G (also called either thefree energy or Gibbs energy) describes both the directionin which a chemical reaction will tend to proceedand the concentrations of reactants and products thatwill be present at equilibrium. ∆G for a chemical reactionequals the sum of the free energies of formation ofthe reaction products ∆G P minus the sum of the freeenergies of formation of the substrates ∆G S . ∆G 0 denotesthe change in free energy that accompanies transitionfrom the standard state, one-molar concentrationsof substrates and products, to equilibrium. A more usefulbiochemical term is ∆G 0′ , which defines ∆G 0 at astandard state of 10 −7 M protons, pH 7.0 (Chapter 10).If the free energy of the products is lower than that ofthe substrates, the signs of ∆G 0 and ∆G 0′ will be negative,indicating that the reaction as written is favored inthe direction left to right. Such reactions are referred toas spontaneous. The sign and the magnitude of thefree energy change determine how far the reaction willproceed. Equation (3)—∆G 0 =− RTln Keq—illustrates the relationship between the equilibriumconstant K eq and ∆G 0 , where R is the gas constant (1.98cal/mol/°K or 8.31 J/mol/°K) and T is the absolutetemperature in degrees Kelvin. K eq is equal to the productof the concentrations of the reaction products, eachraised to the power of their stoichiometry, divided bythe product of the substrates, each raised to the powerof their stoichiometry.(2)(3)