J. Phys. Chem. B 2009, 113, 9995–10000
9995
Protonated Arginine and Protonated Lysine: Hydration and Its Effect on the Stability of Salt-Bridge Structures Bing Gao, Thomas Wyttenbach, and Michael T. Bowers* Department of Chemistry and Biochemistry, UniVersity of California, Santa Barbara, California 93106-9510 ReceiVed: April 9, 2009; ReVised Manuscript ReceiVed: May 27, 2009
Using a mass spectrometer equipped with a drift cell, water binding energies of protonated arginine (ArgH+) and protonated lysine (LysH+) were determined in equilibrium experiments and supplementary calculations at the B3LYP/6-311++G** level of theory. The binding energy of the first water molecule was measured to be 10.3 and 10.9 kcal/mol for ArgH+ and LysH+, respectively. Water binding energies decrease with increasing degree of hydration reaching values of 6-7 kcal/mol for the fourth and fifth water molecule. Theory reproduces this trend of decreasing binding energies correctly and theoretical water binding energies agree with experiment quantitatively within 2 kcal/mol. Lowest-energy theoretical structures of ArgH+ and LysH+ are characterized by protonated side chains and neutral R-amino and carboxyl groups which form intramolecular hydrogen bonds to the ionic group (charge solvation or CS structures). The salt bridge (SB) structures with two cationic groups (side chain and R-amine) and one anionic group (carboxyl) are 13.1 and 9.3 kcal/mol higher in energy for ArgH+ and LysH+, respectively. Theory indicated that the first water molecule binds to the ionic group of the CS structures of ArgH+ and LysH+. With increasing degree of hydration intramolecular interactions are replaced one by one with water bridges with water inserted into the intramolecular hydrogen bonds. Whereas the global minima of ArgH+ · (H2O)n and LysH+ · (H2O)n, n < 7, were calculated to represent CS structures, 7-fold hydrated CS and SB structures, ArgH+ · (H2O)7 and LysH+ · (H2O)7, are nearly isoenergetic (within 0) ArgH+ and LysH+, and Calculated Sequential Hydration Enthalpies ∆Hn0a relative energy (kcal/mol) n
CS
ArgH+ · (H2O)n 0 1 2 3 4 5 6 7 8 LysH+ · (H2O)n 0 1 2 3 4 5 6 7
0 0 0 0 0 0 0 0 1.4 0 0 0 0 0 0 0 0.5
(CS2)b
(3.2) (3.0) (1.7) (0.1) (1.0) (1.8)
SB
-∆Hn0 (kcal/mol)
13.1 10.8 7.6 6.7 4.2 5.6 1.2 0.2 0 9.3 6.7 4.7 2.1 2.5 3.0 4.3 0
s 11.1 10.6 10.4 8.3 7.7 7.1 5.9 5.9 s 11.1 10.2 8.0 7.5 7.2 7.1 6.2
a All values include corrections for BSSE and zero point energies. ∆Hn0 values include thermal energy (298 K) and they are based on the enthalpy difference between the lowest-energy MH+ · (H2O)n and MH+ · (H2O)n-1 structure. b Values in parentheses are energies of R-amine protonated lysine structures (CS2) relative to the lowest-energy structure.
Figure 3. Lowest-energy charge solvation (CS and CS1) and salt bridge (SB) structures of dehydrated ArgH+ and LysH+. The CS2 structure shown for LysH+ corresponds to an energetically less favorable charge solvation structure with protonation on the R-amine.
Why is the lysine salt bridge more stable than the arginine salt bridge with respect to their CS structures? A glance at Figure 3 does not reveal an obvious reason. Therefore we calculated the energy of the structure with fully extended side chain for each MH+ system (shown in Figure S6 in Supporting Information) as a common reference. This calculation indicates that the optimum LysH+ CS structure receives more charge solvation stabilization energy upon folding (-20 kcal/mol) than the optimum ArgH+ CS structure (-15 kcal/mol). The relative stabilization energies on folding of the two optimum SB structures are also very different with -11 and -1 kcal/mol for LysH+ and ArgH+, respectively. Hence, since the arginine and lysine side chains exhibit a similar degree of flexibility, it appears that electrostatics drives the energetics. This is in line with previous work46 on alkali ion cationized small amino acids which has shown that the size of the charge center, the alkali ion in that work, is by far the most important factor determining the energetics of various structures. In the systems considered
Figure 4. Lowest-energy structures of ArgH+ · (H2O)n, n ) 1-5.
Figure 5. Lowest-energy structures of LysH+ · (H2O)n, n ) 1-5. Two isoenergetic structures are shown for LysH+ · (H2O)3, one with protonation on the R-amine and one with protonation on the ε-amine. For LysH+ · (H2O)5 there exists an isoenergetic (0.1 kcal/mol) isomer with identical lysine conformation and identical water bridges as shown here, but with a fifth water molecule bound to the ammonium group instead of the carboxyl group.
here, the tightly focused charge center in LysH+ affects both the charge solvation energy in the CS structure and to an even larger degree the salt bridge interaction energy in the SB structure compared to the ArgH+ system. This is true despite the fact that the ArgH+ SB structure has two side chaincarboxylate hydrogen bonds and LysH+ only one. Given the strong influence of electrostatics, it could in fact be that the SB structure is more competitive in LysH+ than in ArgH+, not because the LysH+ SB structure is particularly stable but because the LysH+ CS structure is not particularly optimal. Figure 3 indicates that only one of the two NH3+ · · · OdC and NH3+ · · · NH2 hydrogen bonds in the LysH+ CS1 structure can be optimal in any given structure, never both simultaneously. In contrast, the guanidinium geometry is in a better position to satisfy geometrical requirements for both hydrogen bonds simultaneously, one to OdC and one to NH2. The optimum singly hydrated ArgH+ · H2O and LysH+ · H2O structures are shown in Figures 4 and 5. It can be seen that H2O simply adds to the ionic site of the optimum dehydrated structure in both cases. The calculated water binding energies agree with the experimental values within
