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B6. Motion and Enzyme Catalysis

  • Page ID
    5540
  • [ "article:topic", "authorname:jjakubowskih" ]

    Benkovic and Hammes-Schiffer have offered an in-depth review of methods used by enzymes in catalysis. They divided their analysis into a biochemical and theoretical frameworks. From a biochemical viewpoint, enzyme catalysis was thought to result from binding of substrate followed by activation of the substrate by the enzyme. The substrate might bind in a particularly reactive conformation in a lock-and-key fashion, or be distorted by the enzyme on binding, making it more susceptible to bond making/breaking. In an energy diagram for the latter case, the bound distorted substrate has higher energy than the free substrate, and is an example of substrate (ground state) destabilization. With the theoretical work of Pauling and development of catalytic antibodies by Lerner, the idea than enzyme functions by binding the transition state with higher affinity than the substrate became accepted. Evidence for tighter transition state binding was found for serine proteases (through stabilization of the tetrahedral oxyaninon intermediate) and lysozyme (through stabilization of the developing oxocarbenium ion (positive charge) by adjacent Asp and Glu residues (both of which are described above). Quantitative increases in catalytic hydrolysis of phosphoesters and diesters of 1015 to 1017 by some phosphatases and nucleases over the uncatalyzed, spontaneous hydrolysis in aqueous solutions are known. Assuming that the ratio of the catalyzed rate at low S (kcat/Km with units of M-1s-1) versus the noncatalyzed rate (pseudo first order with using of s-1) gives a measure of the effective concentration of substrate in the noncatalyzed rate that would give the same rate as the catalyzed rate, effective concentrations reach as high as 1024 for enzymes like orotic acid decarboxylase. What is the source of this catalytic power? Much evidence has been accrued to partition catalysis into contributions from general acid/base catalysis, nucleophilic and electrostatic catalysis, as well as binding effects that enhance catalysis. Solvent effects also show that for some reactions, solvation slows the reaction, implying that desolvation of the substrate during substrate-enzyme binding affects the reaction rate. Remember that the active site in the E-S complex is devoid of most water molecules that are present in the free enzyme. Finally the enzyme may have the correct 3D framework that positions all side chains involved in binding in catalysis in optimal positions, which may have the effect of selecting only catalytically-competent conformations of the substrate (i.e. those the most resemble the transition state). These contributions suggest that movement of critical residues in the active site is required, which positions the substrate and catalytic residues in optimal positions for catalysis. The kinds of motions (which can be modeled in molecular dynamics simulations) are described in the table below.

    Table: Time Scales of Dynamic Events in Enzyme Catalysis
    Motion Approx. Time Scale - log(s)
    bond vibration -14 to -13
    proton transfer -12
    hydrogen bonding -12 to -11
    elastic vibration of globular region -12 to -11
    sugar repuckering -12 to -9
    rotation of side chains at surface -11 to -10
    torsional libration of buried group -11 to -9
    hinge bending at domain interfaces -11 to -7
    water structure reorganization -8
    helix breakdown/formation -8 to -7
    allosteric transitions -5 to 0
    local denaturation -5 to 1
    rotation of medium-sized interior sidechains -4 to 0

    from Benkovic, S and Hammes-Schiffer, S. Science. 301, pg 1197 (2003)

    Warshel argues that electrostatic stabilization of the transition state is the predominate mechanism for tighter binding of the transition state than the substrate to the enzyme. Hydrophobic interactions would facilitate proper orientation of the substrate/transition state in the active site pocket, which is preorganized in effect by the structure of the enzyme. Other have argued that quantum mechanical tunneling (of hydrides, protons, or electrons) is a main mechanism to surmount energetic barriers in enzyme-catalyzed reactions. Using theoretical and molecular dynamic simulations, Gao et al argue that tunneling, a way to overcome classical energy barriers, plays a much smaller role that reducing the barrier height (activation energy). Warshel notes that tunneling effects can also occur in solution in the absence of enzyme, making it all the more important to compare reactions mechanisms in the presence and absence of enzyme. Yet proteins motions that move catalytically important side chains toward the substrate/transition state may allow both preferential electrostatic stabilization of the transition state and quantum mechanical tunneling. Yet another idea states that enzymes must create new bonds to substrate/transition state that could not be available in the absence of the enzyme. One such type of bond is a low-barrier hydrogen bond, in which a hydrogen on an electronegative atom is involved in strong H bonds to two different hydrogen bond acceptors simultaneously. Finally, substrate may adopt near-attack conformations (NAC), which more approximate the bound transition state, and it is the binding of the NAC to the enzyme, not lowering the activation energy, that contributes to the enzyme catalyzed rate enhancement.