Allostery vs. "allokairy".

A hallmark feature of biological systems is that they are tightly regulated. Whether it is turning genes on and off, controlling cell division, or tuning the activity of enzymes, nature has evolved an intricate array of regulatory measures to ensure that systems can optimally respond to the myriad of environmental queues that determine everything from cell fate to survival. Most often the tuning of an enzyme uses a phenomenon known as allostery, whereby the binding of substrate to one enzyme molecule is coupled to the binding of another molecule. The end result is that binding at one site can influence subsequent binding events at other sites. Thus, the term “allostery,” which is derived from the Greek allos meaning “other” and stereos meaning “structure,” describes the ability of biological molecules to transmit the effects of binding spatially through the protein to other sites. The association of oxygen with tetrameric hemoglobin is the prototypical example (1), and indeed almost every enzyme (2) is allosterically controlled by some ligand. However, is the coupling of spatially distinct events the only way to regulate function? In PNAS, Whittington et al. (3) demonstrate how regulation can arise not only by transmitting binding information spatially but also temporally. This mode of regulation forces a reconsideration of the strategies nature has at its disposal to tune biological systems. For an enzyme to be tunable, it is well appreciated that there must exist at least two forms of the molecule, one with a high affinity for substrate and the other with low affinity (Fig. 1A). If the relative fraction of molecules occupying the highand low-affinity states can be adjusted, the enzyme can be mostly unbound under one set of conditions (i.e., more molecules in the low-affinity state), and mostly bound under another (i.e., more molecules in the high-affinity state), thus making the activity tunable. The question is, What strategies will endow an enzyme with such tunability? For decades, since the existence of regulation was first uncovered (4), the vast majority of regulatory mechanisms relied on the coupling between different sites. This phenomenon is rooted in the very simple principle that if there is a difference in binding affinity between two states, the addition of substrate will stabilize (i.e., make more probable) the state that binds with higher affinity. So, how did nature use this principle to regulate function? By evolving so that the functional unit is an oligomer (i.e., a dimer, trimer, or tetramer, etc.) with all of the copies of the functional units being forced to convert from the lowto high-affinity states together, any stabilization of the high-affinity state caused by substrate binding will be propagated to all other copies. Thus, under low-substrate conditions, all of the oligomers would be in the low-affinity state (Fig. 1B, Left). Addition of substrate would have the dual effect of binding to some of the sites (Fig. 1B, Middle) and also modulating the relative amounts of molecules in the highand low-affinity states, transforming the remaining empty sites into high-affinity sites. The more substrate that is added, the more sites that become bound and the higher the affinity becomes (Fig. 1B, Right). This principle, the coupling of binding at two spatially separated sites, is at the heart of allostery, and up until now was believed to underlie almost all regulation. In the late 1960s, however, it was becoming increasingly apparent that the rate of isomerization of an enzyme could also influence regulation, with some enzymes showing a slow response to changes in the concentrations of substrate (5). It was proposed that such “hysteretic” enzymes should display properties that resemble the classic behavior of oligomeric allosteric systems (5–10). However, unlike those allosteric systems, the origin of the effect should not originate from the through-space coupling of different binding sites. The effect should be entirely kinetic, depending instead on a competition between two processes, the binding of the enzyme to more substrate and the relaxation of the enzyme back to its original low-affinity state. For such a system, when substrate concentration is low (Fig. 1C, Left), product release is followed by nearly all of the enzymes relaxing back into the low-affinity state before encountering another substrate molecule. Fig. 1. Allosteric and allokairic regulation. (A) Tunability requires that an enzyme can populate at least two states, depicted here as low-affinity (L) and high-affinity (H) with the ligand represented as the blue oval. (B) In allostery, binding and conformational change are coupled. Binding substrate to one molecule of a dimer, for example, converts both to the high-affinity H state, increasing the binding affinity. (C ) In allokairy, the enzyme is in the H state after turnover (upper right) and relaxes back to the L state (lower left) determined by the time between turnover and binding, which depends on substrate concentration ([S]). (D) Allostery (two-site dimer, light gray; four-site tetramer, dark gray) and allokairy (red dashed lines) both lead to S-shaped curves when activity is plotted against substrate concentration. The curves for a fully L state or H state are shown in blue and green (top and bottom). Although both regulation mechanisms produce a sigmoidal change in activity, transitioning from the L state at low [S] to the H state at high [S], cooperativity in allokairy is more tunable (gray shading).