Poutsma has followed up on the work he reported earlier in collaboration with Kass concerning the gas-phase acidity of the amino acids.¹ Their previous work reported on cysteine,² with the unusual result that the thiol group is more acidic than the carboxylic acid group. (I blogged on this a previous post.) Now, he reports the experimental and DFT acidities of all 20 amino acids, shown in Table 1. The experiments were done using the kinetic method in a quadrupole ion trap with electrospray ionization. The computations were performed at B3LYP/6-311++G**//B3LYP/6-31+G*, following some MM searching to identify low-lying conformations. The computed acidities were obtained relative to acetic acid, i.e. R-CH₂COOH + OAc^– → R-CH₂COO^– +HOAc.

Table 1. Relative acidities (kJ mol^-1) of the amino acids¹

The computed values are in very good agreement with the experimental values. The amino acids are ordered in increasing acidity in Table 1. The order predicted by experiment and DFT are quite close, and the disagreements are well within the error bar of the experiment.

Similar to the result for cysteine, tyrosine also displays unusual acidity. The alcohol proton is more acidic than the carboxylic acid proton. The structures of tyrosine, and its two conjugate
bases, one from loss of the phenolic proton and the other from loss of the carboxylic acid proton are shown in Figure 1. The stability of the tyrosine conjugate base from loss of the phenolic
hydrogen arises from both the stability of phenoxide and the internal hydrogen bond from the carboxylic acid proton to the amine. This is different that in the cysteine case, the thiolate anion is stabilized by an internal hydrogen bond from the carboylic acid group (see Figure 2c here).

tyrosine
Tyrosine conjugate base (loss of phenolic hydrogen)	Tyrosine conjugate base (loss of carboxylate hydrogen)

Figure 1. B3LYP/6-31G* optimized structures of tyrosine and its conjugate bases.¹

References

(1) Jones, C. M.; Bernier, M.; Carson, E.; Colyer, K. E.; Metz, R.; Pawlow, A.; Wischow, E. D.; Webb, I.; Andriole, E. J.; Poutsma, J. C., "Gas-Phase Acidities of the 20 Protein Amino Acids," Int. J. Mass Spectrom. 2007, 267, 54-62, DOI: 10.1016/j.ijms.2007.02.018.

(2) Tian, Z.; Pawlow, A.; Poutsma, J. C.; Kass, S. R., "Are Carboxyl Groups the Most Acidic Sites in Amino Acids? Gas-Phase Acidity, H/D Exchange Experiments, and Computations on Cysteine and Its Conjugate Base," J. Am. Chem. Soc. 2007, 129, 5403-5407, DOI: 10.1021/ja0666194.

InChI

Tyrosine: InChI=1/C9H11NO3/c10-8(9(12)13)5-6-1-3-7(11)4-2-6/h1-4,8,11H,5,10H2,(H,12,13)/t8-/m0/s1

Kass has once again uncovered a simple system that challenges our notions of basic chemical concepts. It is a well accepted notion that the most acidic proton of all of the amino acids is the carboxylic acid one. However, acidities are strongly influenced by the solvent, and the absence of solvent in the gas phase can dramatically alter things.

Kass and co-workers examined the gas-phase acidity of cysteine with computational and
experimental techniques.¹ The lowest energy conformer of cysteine is 1a, characterized by having three intramolecular hydrogen bonds (Figure 1). The next lowest conformer, 1b, has only two intramolecular hydrogen bonds and is 1.5 kcal mol^-1 higher in energy at G3B3.

1a xyz	1b xyz

Figure 1. B3LYP/aug-cc-pVDZ optimized structures of cysteine 1.¹

They optimized a number of different configurations of the conjugate base of cysteine: two conformers from the loss of the carboxylate proton (2a and 2b), two conformers from the loss of the thiol proton (2c and 2d), and one conformer from the loss of the thiol proton of the zwitterion (2e). These structures are shown in Figure 2 along with their relative energies. All of these structures possess two intramolecular hydrogen bonds.

2a (3.1) xyz	2b (3.4) xyz
2c (0.0) xyz	2d (5.1) xyz
2e (10.1) xyz

Figure 2. B3LYP/aug-cc-pVDZ optimized structures of the conjugate base of cysteine 2. Relative energies (kcal mol^-1) in parenthesis computed at G3B3.¹

The gas phase acidity of carboxylic acids is greater than thiols; the deprotonation energy of propanoic acid (CH₃CH₂CO₂H) is 347.7 kcal mol^-1 at G3B3 (347.2 expt.²), about 6 kcal mol^-1 less than that of ethanethiol (CH₂CH₂SH: 355.0 at G3B3 and 354.2 expt.²). However, the computations indicate that 2c is the lowest energy structure of deprotonated cysteine, and 2c comes about by loss of the thiol proton! Te lowest energy cysteine conjugate base from loss of the carboxylate proton is 1a, which is 3.1 kcal mol^-1 higher in energy. Apparently, the hydrogen bonding network in 2c is quite favorable, able to make up for the inherent favorability of a carboxylate over a thiolate anion.

The G3B3 computed deprotonation energy of cysteine is 333.3 kcal mol^-1 (for removal of the thiol proton). Kass determined the deprotonation energy of cysteine using a kinetic and a thermodynamic method. The kinetic method gives a value of 332.9 ± 3.3 kcal mol^-1­, while the thermodynamic method gives 334.4 ± 3.3 kcal mol^-1­. These are in fine agreement with the computed value.

This study ably demonstrates the dramatic role that solvent can play in determining molecular properties. Kass titled the article “Are carboxyl groups the most acidic sites in amino acids?” and answers with “no” – in the gas phase the thiol group is more acidic. He ends the article with an indication that the alcohol of tyrosine may be competitive in acidity with its carboxylic group, too.

References

(1) Tian, Z.; Pawlow, A.; Poutsma, J. C.; Kass, S. R., "Are Carboxyl Groups the Most Acidic Sites in Amino Acids? Gas-Phase Acidity, H/D Exchange Experiments, and Computations on Cysteine and Its Conjugate Base," J. Am. Chem. Soc., 2007, 129, 5403-5407, DOI: 10.1021/ja0666194.

(2) NIST, NIST Chemistry WebBook, 2005, http://webbook.nist.gov/.

InChIs

1: InChI=1/C3H7NO2S/c4-2(1-7)3(5)6/h2,7H,1,4H2,(H,5,6)