2 Структура и функция белка (1160071), страница 29
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This is illustrated by the energetic "hill" in Figures 8-3 and 8-4.To undergo reaction, the molecules must overcome this barrier andtherefore must be raised to a higher energy level. At the top of the energyhill is a point at which decay to the S or P state is equally probable (itis downhill either way). This is called the transition state. The transition state is not a chemical species with any significant stability andshould not be confused with a reaction intermediate. It is simply afleeting molecular moment in which events such as bond breakage,bond formation, and charge development have proceeded to the precisepoint at which a collapse to either substrate or product is equallylikely.
The difference between the energy levels of the ground state andthe transition state is called the activation energy (AG*). The rate ofa reaction reflects this activation energy; a higher activation energycorresponds to a slower reaction. Reaction rates can be increased byraising the temperature, thereby increasing the number of moleculeswith sufficient energy to overcome this energy barrier. AlternativelyChapter 8 Enzymesthe activation energy can be lowered by adding a catalyst (Fig. 8-4).Catalysts enhance reaction rates by lowering activation energies.Enzymes are no exception to the rule that catalysts do not affectreaction equilibria.
The bidirectional arrows in Equation 8-1 makethis point: any enzyme that catalyzes the reaction S —» P also catalyzesthe reaction P —» S. Its only role is to accelerate the interconversion ofS and P. The enzyme is not used up in the process, and the equilibriumpoint is unaffected. However, the reaction reaches equilibrium muchfaster when the appropriate enzyme is present because the rate of thereaction is increased.This general principle can be illustrated by considering the reaction of glucose and O2 to form CO2 and H2O. This reaction has a verylarge and negative AG°', and at equilibrium the amount of glucosepresent is negligible. Glucose, however, is a stable compound, and itcan be combined in a container with O2 almost indefinitely withoutreacting. Its stability reflects a high activation energy for reaction. Incells, glucose is broken down in the presence of O2 to CO2 and H2O in apathway of reactions catalyzed by enzymes.
These enzymes not onlyaccelerate the reactions, they organize and control them so that muchof the energy released in this process is recovered in other forms andmade available to the cell for other tasks. This is the primary energyyielding pathway for cells (Chapters 14 and 18), and these enzymesallow it to occur on a time scale that is useful to the cells.In practice, any reaction may have several steps involving the formation and decay of transient chemical species called reaction intermediates. When the S ^ P reaction is catalyzed by an enzyme, the ESand EP complexes are intermediates (Eqn 8-1); they occupy valleys inthe reaction coordinate diagram (Fig.
8-4). When several steps occurin a reaction, the overall rate is determined by the step (or steps) withthe highest activation energy; this is called the rate-limiting step. Ina simple case the rate-limiting step is the highest-energy point in thediagram for interconversion of S and P (Fig.
8-4). In practice, the ratelimiting step can vary with reaction conditions, and for many enzymesseveral steps may have similar activation energies, which means theyare all partially rate-limiting.As described in Chapter 1, activation energies are energetic barriers to chemical reactions; these barriers are crucial to life itself. Thestability of a molecule increases with the height of its activation barrier.
Without such energetic barriers, complex macromolecules wouldrevert spontaneously to much simpler molecular forms. The complexand highly ordered structures and metabolic processes in every cellcould not exist. Enzymes have evolved to lower activation energiesselectively for reactions that are needed for cell survival.Reaction Rates and Equilibria Have PreciseThermodynamic DefinitionsReaction equilibria are inextricably linked to AG°' and reaction ratesare linked to AG¥.
A basic introduction to these thermodynamic relationships is the next step in understanding how enzymes work.As introduced in Chapter 4, an equilibrium such as S ^ P is described by an equilibrium constant, Keq. Under the standard conditions used to compare biochemical processes, an equilibrium constantis denoted K(eqK '=1*1eqIS](8-2)203Transition state ($)Reaction coordinateFigure 8-4 Reaction coordinate diagram comparing the enzyme-catalyzed and uncatalyzed reactionsS —> P.
The ES and EP intermediates occupy minima in the energetic progress curve of the enzymecatalyzed reaction. The terms AG5ncat and AGjat correspond to the activation energies for the uncatalyzed and catalyzed reactions, respectively. Theactivation energy for the overall process is lowerwhen the enzyme catalyzes the reaction.204Part II Structure and CatalysisFrom thermodynamics, the relationship between Keq' and AG°' can bedescribed by the expression^GO' = -RT\nK,tTable 8-4 The relationshipbetween Keq' and AG°' (seeEqn 8-3)K'io- 610~5io- 410" 310" 2lO" 11IO1IO2IO3AG°' (kJ/mol)34.228.522.817.111.45.70.0where R is the gas constant (8.315 J/mol • K) and T is the absolutetemperature (298 K).
This expression will be developed and discussedin more detail in Chapter 13. The important point here is that theequilibrium constant is a direct reflection of the overall standard freeenergy change in the reaction (Table 8-4). A large negative value forAG°' reflects a favorable reaction equilibrium, but as already notedthis does not mean the reaction will proceed at a rapid rate.The rate of any reaction is determined by the concentration of thereactant (or reactants) and by a rate constant, usually denoted by thesymbol k. For the unimolecular reaction S —» P, the rate or velocity ofthe reaction, V, representing the amount of S that has reacted per unittime, is expressed by a rate law:V = k[S]-5.7-11.4-17.1(8-3)(8-4)In this reaction, the rate depends only on the concentration of S.
This iscalled a first-order reaction. The factor k is a proportionality constantthat reflects the probability of reaction under a given set of conditions(pH, temperature, etc.). Here, k is a first-order rate constant and hasunits of reciprocal time (e.g., s"1). If a first-order reaction has a rateconstant k of 0.03 s"1, this may be interpreted (qualitatively) to meanthat 3% of the available S will be converted to P in 1 s. A reaction witha rate constant of 2,000 s" 1 will be over in a small fraction of a second.If the reaction rate depends on the concentration of two different compounds, or if two molecules of the same compound react, the reaction issecond order and k is a second-order rate constant (with the unitsM~ 1 S~ 1 ).
The rate law has the formV=«SJ[S 2 ](8-5)From transition-state theory, an expression can be derived thatrelates the magnitude of a rate constant to the activation energy:kTh(8-6)where k is the Boltzmann constant and h is Planck's constant. Theimportant point here is that the relationship between the rate constant, k, and the activation energy, AG*, is inverse and exponential. Insimplified terms, this is the basis for the statement that a lower activation energy means a higher reaction rate, and vice versa.Now we turn from what enzymes do to how they do it.A Few Principles Explain the Catalytic Powerand Specificity of EnzymesTable 8-5 Some rateenhancements producedby enzymesCarbonic anhydrasePhosphoglucomutaseSuccinyl-CoA transferaseUreaseIO 7IO12IO1310 14Enzymes are extraordinary catalysts.
The rate enhancements broughtabout by enzymes are often in the range of 7 to 14 orders of magnitude(Table 8-5). Enzymes are also very specific, readily discriminating between substrates with quite similar structures. How can these enormous and highly selective rate enhancements be explained? Wheredoes the energy come from to provide a dramatic lowering of the activation energies for specific reactions?Chapter 8 Enzymes205Part of the explanation for enzyme action lies in well-studiedchemical reactions that take place between a substrate and enzymefunctional groups (specific amino acid side chains, metal ions, and coenzymes). Catalytic functional groups on enzymes can interact transiently with a substrate and activate it for reaction.
In many cases,these groups lower the activation energy (and thereby accelerate thereaction) by providing a lower-energy reaction path. Common types ofenzymatic catalysis are outlined later in this chapter.Catalytic functional groups, however, are not the only contributorto enzymatic catalysis. The energy required to lower activation energies is generally derived from weak, noncovalent interactions betweenthe substrate and the enzyme.
The factor that really sets enzymesapart from most nonenzymatic catalysts is the formation of a specificES complex. The interaction between substrate and enzyme in thiscomplex is mediated by the same forces that stabilize protein structure, including hydrogen bonds and hydrophobic, ionic, and van derWaals interactions (Chapter 7). Formation of each weak interaction inthe ES complex is accompanied by a small release of free energy thatprovides a degree of stability to the interaction.
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