Week #4 (Due Sunday, Feb. 8)

An amino acid can be present in two forms: the un-ionized form, H2NCHRCO2H, and the Zwitterion, +H3NCHRCO2-. A zwitterions is a molecule which contains ions but which as a whole is neutral.

You will explore the properties and the relative stability of the two forms in the case of the simplest amino acid glycine. A series of calculations will be performed on each structure. You will find the following sequence of operations to be useful after drawing the structure: save the file using Save As, define the calculation, and perform it using Submit. This approach will guarantee that you will end up with a Spartan file for each step of the calculations.

I) Calculations on glycine in the Zwitterion form, +H3NCH2CO2-.

A) Draw the structure of your amino acid using the Peptide feature of the Build menu. Use the Terminate feature to properly define the charges at the termini.

B) Minimize the energy of the structure using Molecular Mechanics with the Merck Molecular Force Field. The Calculate Field of the Setup menu should be set to Equilibrium Conformer. Comment on the conformer produced by the minimization. Why this particular conformer?

This structure maximizes the interaction/attraction of the two charged ends of the zwitterion.

Measure the bond lengths, bond angles, and torsional angles.

C) Re-minimize the structure obtained in step (B) using a semi-empirical quantum mechanical method (Calculate à Semi-Empirical) and an AM1 Hamiltonian. Measure the bond lengths, bond angles, and torsional angles of the resulting structure. Compare these values to the structure from part B and literature values.

The crystal structure for the three forms of glycine (+H3NCH2COO-, H2NCH2COO-, and H3NCH2COOH ) was extracted from the Cambridge Structural Database and is in the file ‘glycine.pdb’. You can use Spartan to open the file.

The most notable difference is the hydrogen and oxygen are no longer aligned in the C-C-N plane as in the calculated structures. This illustrates the effects of environment (e.g. sulfate ion and crystal packing) on the equilibrium structure.

Such drastic conformational difference leaves little allowance for method comparison. Examining individual moieties AM1 does no better reproducing C-O bond lengths or C-H bond lengths relative to MMFF although slightly better agreement for the OCO,OCC and CCN angles exists, it is marginal. The MMFF parameter set includes glycine dipeptide so it should not be too suprising it performs satisfactorily.

II) Calculations on glycine in the un-ionized form, H2NCH2CO2H.

A) Draw the structure of your amino acid using the Peptide feature of the Build menu. Use the Terminate feature to properly define the terminal groups.

B) The unionized species is conformationally more flexible. Perform a search of low energy conformers. The Calculate field of the Setup menu should be set to Conformer Distribution. After completing the calculation load (File à Open … should find file with .Conformer suffix) the spreadsheet created by the process and examine the low-energy conformers.

To display the molecules and energies:

1) Choose Display à Spreadsheet

2) Add à E à kcal/mol à OK

3) Energies of the conformers should appear in the spreadsheet

To graph the energies and molecules

1) Choose Display à Plot

2) In window choose Molecule for the X-axis and E for the Y-axis à OK

Consider the most direct conversion of the Zwitterion to the unionized form. The hydrogen on the –NH3+ that is closest to a carboxylate oxygen is transferred as a proton to that oxygen. Examine several low-energy conformers and find the conformer that matches the description of this species. Is it the conformer with the lowest energy?

Examine the energies of other low-energy conformers and compare and contrast their structures.

MMFF: Yes. The next highest energy conformer is due to rotation of the carboxyl group 180 such that the OH group faces away from the NH2 leading to less (according to MMFF) H-bonding type interactions as the carbonyl oxygen is ~2.6 Angstroms from the NH2 hydrogens compared to ~1.8 Angstrom distance between the hydroxyl hydrogen and nitrogen in the lowest energy structure. The next highest energy structure represents a 180 rotation of the hydroxyl group from the lowest energy conformer.

Thus these two conformers correspond to cis (OH and NH2 on same side of CC bond) and trans (opposite sides), the cis being more sterically crowded. Although at room temperature the two forms would be present in comparable amounts. The next two conformers are similar to 2 and 3 except the amide group is rotated ninety degrees. Finally the highest energy group represents a 180 rotation of the hydroxyl group in conformer two introducing additional strain interactions with the hydrogens on the methyl carbon with the loss of the carbonyl oxygen – hydroxyl hydrogen stabilization interaction.

AM1: No. The two lowest energy structures from AM1 have the hydroxyl hydrogen pointed away from the amide group contrary to what one expects intuitively and based on the MMFF calculations. Looking at the crystallographic file the neutral moiety is closer to the AM1 result with the amide group rotated 90 (perhaps due to the environment). Experiments¹ with Hartree-Fock calculations and Moller-Plesset ab initio methods (using the 6-31G** basis set) confirm the lowest two non-zwitterionic conformers predicted by AM1. The energy difference between the neutral and zwitterionic forms is 42.38 kcal/mol compared to the MP2 result of 22.57 kcal/mol. This indicates the MMFF model underestimates the intramolecular hydrogen bonding between the carboxyl oxygen and amide hydrogens and/or overestimates the H-bonding between nitrogen and hydrogen. This is somewhat suprising considering the MMFF force field did include di-glycine in its parameterization.

The next highest structure is simply a 180 rotation of carboxylate group and hence higher in energy due to less favorable interactions between hydroxyl and amide group. The next group represents a 90 rotation of amide group bringing several hydrogens into the higher energy gauche-like geometry. The next higher energy conformer represents a 90 rotation of the hydroxyl group of the lowest energy conformer resulting in non-favorable interactions with the CH2 hydrogens. The next highest represents rotation of the amide and hydroxyl group. Finally the next four structures,comparable in energy, are representative of the transition between the neutral and zqitterionic form with the hydroxyl hydrogen indirectly facing the amide nitrogen.

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¹ Kwon, O. Y. et al. Bull. Korean Chem Soc., 16(5), 410 (1995).