Correlation between Asp115 and the Retinal Schiff Base

β-ionone ring of the retinal on its cytoplasmic side. As stated in the previous subsection, Asp115 is not a proton donor or acceptor during the proton transfer in BR. It is, however, implicated in regulating the proton transfer. Asp115 is protonated in all intermediate

5.5. Correlation of the Protonation Behavior 109

0 2 4 6 8 10 12 14

pH

-1 -0.5 0 0.5 1

c ij

bR:1c3w bR:1f50 L:1vjm.b M:1f4z M:1kg8 N:1p8u

Figure 5.21. Correlation between Asp115 and the retinal Schiff base. The coeffi-cient of the correlation between Asp115 and the retinal Schiff base is plotted against pH for six BR structures.

protonation states of BR. The retinal Schiff base is protonated in the [bR], [N] and [O]

protonation state and deprotonated in the [M1] and [M2] state. The photocycle and its proton transfer steps are depicted in Figure 5.2 and the intermediate protonation states are listed in Table 5.1.

The correlation between Asp115 and the retinal Schiff base is shown in Figure 5.21 for six BR structures. No significant correlation can be observed for bR:1c3w, bR:1f50 or L:1vjm.b. In the two M structures, M:1f4z and M:1kg8, and the N structure N:1p8u, however, the protonation behavior of Asp115 and the retinal Schiff base is negatively correlated. In the following subsections, the correlation behavior is discussed in detail for the individual structures.

BR STRUCTURES

For both bR:1c3w and bR:1f50, the correlation coefficient is 0 at pH<7.5. In the pH range from 7.5 to 14, the correlation coefficient takes values between 0 and -0.1. As shown in Figure 5.10 a, Asp115 is fully protonated in the pH range from 0 to 7.5 in both structures. Thereafter, Asp115 deprotonates. At pH = 14, the protonation probability is less than 0.1 in bR:1c3w and about 0.2 in bR:1f50. The protonation probability of the retinal Schiff base is close to 1 over the complete pH range from 0 to 14 for both bR:1c3w and bR:1f50 (cf. Figure 5.11 a). In the pH range from 0 to 7.5, the covariance and the correlation are 0 for both bR state structures due to the absent variance in Asp115.

At higher pH values, when the correlation can be assessed the protonation behavior of Asp115 and the retinal Schiff base is uncorrelated.

L STRUCTURE

For L:1vjm.b, the correlation coefficient is 0. Only at pH>13 at four pH values, an insignificant negative correlation can be observed. As shown in Figure 5.10 c, Asp115 is fully protonated. Only at pH>13, a protonation probability slightly less than 1 can be observed at five pH values. As shown in Figure 5.11 c, at pH<2 the protonation

probability of the retinal Schiff base is close to 1. In the pH range from 2 to 8, the Schiff base deprotonates. At pH>8, the protonation probability is close to 0 or equal to 0. In general, the correlation coefficient is 0 due to the absent variance of Asp115. However, at one of the five pH values, where Asp115 shows variance, the retinal Schiff base is fully deprotonated. Thus, a correlation coefficient unequal to 0 can only be observed at four pH values. The correlation coefficient at the four pH values where both residues show variance indicates that the protonation behavior of Asp115 and the retinal Schiff base is uncorrelated in L:1vjm.b

M STRUCTURES

For M:1f4z and M:1kg8, the correlation of the protonation behavior of Asp115 and the retinal Schiff base is 0 in the pH range from 0 to 6. At pH>6, the correlation decreases.

In the pH range from 10 to 14, Asp115 and the Schiff base show significant negative correlation in both structures. In M:1f4z, the correlation decreases considerably to about -0.7 at pH = 14. In M:1kg8, the correlation coefficient fluctuates between -0.4 and -0.5.

In the pH range from 0 to 6, Asp115 is fully protonated in both M:1f4z and M:1kg8 (cf. Figure 5.10 d). In the pH range from 6 to 10, the protonation probability is close to 1 for both structures. In the pH range from 10 to 14, the probability remains high in M:1kg8, while in M:1f4z the probability decreases to 0.6. As shown in Figure 5.11 d, at pH<4 the protonation probability of the retinal Schiff base is close to 1. In the pH range from 4 to 8, the Schiff base deprotonates. In M:1f4z, the minimum protonation probability is approximately 0.15. At pH = 10, the protonation probability increases up to approximately 0.5 at pH = 14. In M:1kg8, the protonation probability is close to 0 in the pH range from 8 to 14.

In the pH range from 0 to 6, the covariance and the correlation of Asp115 and the retinal Schiff base are 0 due the absent variance of Asp115 in both structures. In the pH range from 6 to 10, when the protonation probability of Asp115 is 0.99 in both structures,i.e., a small variance is observed, the correlation is slightly negative. In the pH range from 10 to 14, Asp115 is partly deprotonated and the Schiff base is partly protonated in M:1f4z.

In contrast in M:1kg8, the protonation probability of Asp115 remains high and that of the Schiff base remains low. The different behavior of Asp115 and the Schiff base in M:1f4z and M:1kg8 results in a more pronounced negative correlation in M:1f4z.

N STRUCTURE

For N:1p8u, the correlation coefficient is 0 in the pH range from 0 to 5. In the pH range from 5 to 11.5, the correlation between Asp115 and the retinal Schiff base de-creases to about -0.8. Thereafter, the coefficient inde-creases to -0.5 at pH = 14. As shown in Figure 5.10 e, Asp115 is fully protonated up to pH = 5. Up to pH = 9, the protonation probability is 0.99. Thereafter, Asp115 slowly deprotonates. At pH = 14, its protonation probability is approximately 0.6. As shown in Figure 5.11 e, at pH<3 the retinal Schiff base is protonated. In the pH range from 3 to 8, the Schiff base deprotonates. At pH = 8, the protonation probability is close to 0. In the pH range from 8 to 14, the probability increases slightly to approximately 0.2. In the pH range from 7 to 14, the protonation behavior of Asp115 and the retinal Schiff base is negatively correlated. Thus, in most

5.5. Correlation of the Protonation Behavior 111 protonation states where Asp115 is protonated, the Schiff base is deprotonated andvice versa.