Volume fluctuation of reaction intermediates determines reactivity

Fig. 6. in Terazima et al, PNAS (2014) shows that a monoactivated protein decamer expands in volume and consequently dissociates, while a polyactivated decamer shrinks in volume and doesn't dissocitate.

 

Proteins are subjected to thermal fluctuations that occur at all length- and timescales, ranging from bond vibrations to large-scale domain rearrangements. The fluctuations affect the volume of the protein and vice versa. The close relationship between protein conformation and protein activity is a well established concept, while the role of fluctuations in protein activity is yet to be elucidated. A recent experimental study on a blue-light photosensor (TePixD) from the thermophile T. elongatus has shown that the flexibility of reaction intermediates is crucial in the overall reaction kinetics.

In solutions and crystals, TePixD proteins form pentameric rings, which further associate to decameric structures. An equilibrium between the pentamers and decamers is impaired by light illumination, upon which the decamers dissociate to pentamers. If the light-source is then removed, the pentamer-decamer equilibrium is restored within 5 s.

Using a pulse laser-based technique, two intermediate states have been identified in the decamer dissociation upon light illumination. These intermediates are named I1 and I2, and they exhibit different volume compressibilities with respect to the ground state. It has been shown that the change in compressibility, which is proportional to change in volume, is positive and about 10% of the compressibility of the protein, meaning that the intermediate species’ volume is larger as compared to the ground state. The volume change is related to different fluctuation levels. It is presumed that the fluctuations are unequally distributed and that the enhancement of the fluctuations at the interface regions of the decamers facilitates the dissociation.

The light illumination causes a conformational change in the immediate vicinity of the photoreactive center, but this center is not near the protein interfaces that are affected by dissociation and association. Evidently, the protein flexibility is changed by the conformational change around the photoactive center, which causes a greater protein flexibility, which in turn increases the volume of the intermediate species and drives protein dissociation.

This is also confirmed by the fact that high-intensity light, which activates more than one protein in the decamer, produces less pentamers because the volume of these species is decreased as well as the protein fluctuations. Thus, there exists an experimental proof that relates higher protein flexibility to enhanced reactivity.