Enzyme dynamics point to stepwise conformational selection in catalysis

Abstract

Recent data increasingly reveal that conformational dynamics are indispensable to enzyme function throughout the catalytic cycle, in substrate recruiting, chemical transformation, and product release. Conformational transitions may involve conformational selection and induced fit, which can be viewed as a special case in the catalytic network. NMR, X-ray crystallography, single-molecule FRET, and simulations clearly demonstrate that the free enzyme dynamics already encompass all the conformations necessary for substrate binding, preorganization, transition-state stabilization, and product release. Conformational selection and substate population shift at each step of the catalytic turnover can accommodate enzyme specificity and efficiency. Within such a framework, entropy can have a larger role in conformational dynamics than in direct energy transfer in dynamically promoted catalysis.

Figure 1

Conformational fluctuations of enzymes lead to ‘catalytic networks’. Eqn (1) is the classical Michaelis–Menten equation. Eqn (2) considers that the conformational ensemble of enzymes allows multiple catalytic reactions to occur in parallel, forming catalytic networks. Eqn (3) illustrates a narrowly defined conformational selection pathway, where only unbound enzyme has multiple conformers while the Michaelis ES and the Product EP complexes are not affected by conformational fluctuations. Eqn (4) is an induced fit mechanism where substrate binding leads to an ES complex with different enzyme conformation.

Figure 2

Conformational dynamics of the binding site for the extrinsic domain of the iron-sulfur protein (ISP-ED) subunit in the cytochrome b (cyt b) subunit of the cyt bc₁ complex is coupled with large motion in ISP-ED (Ref. [18]). MD simulations of the bc1 complex from the R. sphaeroides (Rsbc1) with the bound inhibitor stigmatellin reveal that if no inhibitor exists, the binding pocket opens (B Ma, L Esser, R Nussinov, D Xia, unpublished). The Rsbc1 catalyzes the electron transfer from ubiquinol to cytochrome c and simultaneously pumps protons across the membrane. In the right panel, the simulated nonsymmetrical dimer complex shows that the inhibitor (blue balls) is bound to only to one cyt b subunit (depicted by a red ribbon), while the other cyt b subunit (green ribbon) has an apo binding site. The yellow ribbons are cyt c1 subunits. The ISP subunit with the 2Fe–2S clusters are shown in pink ribbons. The ISP-ED can have a large motion accompanying the electron transfer reaction. In the left panel, two b subunits are superimposed to compare the binding site changes. While the bound subunit (red ribbon) keeps the closed form, the apo subunit (green ribbon) opened up during simulation.

Scheme 1

Multiple conformational selection steps can boost enzyme specificity. This hypothetical scheme illustrates that if there is only 50% specificity for each of the three steps, only 12.5% lead to product release, assuming that the rates of other pathways are slow.