W&M ScholarWorks Undergraduate Honors Theses Theses, Dissertations, & Master Projects 4-2019 Determinations of Proton Affinities of Methylated Cysteine and Serine Homologs Danielle Long Follow this and additional works at: https://scholarworks.wm.edu/honorstheses Part of the Analytical Chemistry Commons, and the Physical Chemistry Commons Recommended Citation Long, Danielle, "Determinations of Proton Affinities of Methylated Cysteine and Serine Homologs" (2019). Undergraduate Honors Theses. Paper 1351. https://scholarworks.wm.edu/honorstheses/1351 This Honors Thesis is brought to you for free and open access by the Theses, Dissertations, & Master Projects at W&M ScholarWorks. It has been accepted for inclusion in Undergraduate Honors Theses by an authorized administrator of W&M ScholarWorks. For more information, please contact [email protected]. Scanned with CamScanner Abstract Non-protein amino acids (NPAAs) are of interest to study for their potential to be incorporated into peptides and proteins, as well as for understanding their structure-energetics relationships. Studying these amino acids’ thermochemical properties such as their acidity and basicity allow for elucidation of the structure and bonding characteristics of these molecules. By examining these molecules in the gas-phase in mass spectrometers, we are able to determine their intrinsic thermodynamic properties without solvation effects. These thermochemical properties, in part, determine the structure, function and bonding of the molecules. This study focused on determining the proton affinities of methylated cysteine and serine homologs by using Cook’s extended kinetic method with orthogonal distance regression analysis in a triple quadrupole mass spectrometer. The NPAAs selected have been shown to mis- incorporate into small peptides, making it of interest to determine how these changes affect the structures and energetics of the peptide.1 The NPAAs studied included α-methylcysteine, L- penicillamine (gem-dimethyl cysteine), α-methylserine, and 3-methylthrenonine (gem-dimethyl serine), and their proton affinities were determined to be 923.7 ± 11 kJ/mol, 925 ± 15 kJ/mol, 932 ± 20 kJ/mol and 924 ± 15 kJ/mol, respectively. These experimental proton affinities are in excellent agreement with Boltmann-weighted computational proton affinities determined for these molecules. All of the methylated homologs had larger proton affinities than their respective protein amino acid, serine and cysteine, which have proton affinities of with proton affinities of 903 kJ/mol and 912 kJ/mol respectively.2 i Table of Contents Chapter 1: Introduction 1 1.1 Acid and Base Chemistry 1 1.2 Previous Non-Protein Amino Acid Studies 2 1.3 Kinetic Method 3 1.4 Quadrupole Mass Spectrometry 7 Chapter 2: Experimental Procedures 10 2.1 Experimentally Determined Proton Affinities 10 2.2 Computationally Determined Proton Affinities 12 Chapter 3: Results and Discussion 13 3.1 Computational Results 14 3.2 α-Methylcysteine Results 16 3.3 L-Penicillamine Results 20 3.4 α-Methylserine Results 24 3.5 3-Methylthreonine Results 28 3.6 Experimental Results Summary 32 Chapter 4: Conclusion and Future Work 34 References 35 ii Figures and Tables Figure 1: Kinetic Method Equation 5 Figure 2: Competitive Dissociation 6 Figure 3: Schematics of Quadrupole Mass Spectrometry 8 Figure 4: α-Methylcysteine 13 Figure 5: L-Penicillamine (dimethyl cysteine) 13 Figure 6: α-Methylserine 13 Figure 7: 3-Methylthreonine (dimethyl serine) 13 Figure 8: Yields of Peptide Produced by Valine, 3-Methylthreonine and L-Penicillamine 14 Table 1: Calculated Proton Affinities 14 Figure 9: Low Energy Conformers of Cysteine Homologs 15 Figure 10: Low Energy Conformers of Serine Homologs 15 Table 2: α-Methylcysteine Experimental Data 16 Figure 11: α-Methylcysteine Plot 1 17 Figure 12: α-Methylcysteine Plot 2 18 Figure 13: α-Methylcysteine Effective Temperature 19 Figure 14: α-Methylcysteine ODR 20 Table 3: L-Penicillamine Experimental Data 20 Figure 15: L-Penicillamine Plot 1 21 Figure 16 L-Penicillamine Plot 2 22 Figure 17: L-Penicillamine Effective Temperature 23 Figure 18: L-Penicillamine ODR 24 Table 4: α-Methylserine Experimental Data 24 iii Figure 19: α-Methylserine Plot 1 25 Figure 20: α-Methylserine Plot 2 26 Figure 21: α-Methylserine Effective Temperature 27 Figure 22: α-Methylserine ODR 28 Table 5: 3-Methylthreonine Experimental Data 28 Figure 23: 3-Methylthreonine Plot 1 29 Figure 24: 3-Methylthreonine Plot 2 30 Figure 253-Methylthreonine Effective Temperature 31 Figure 26: 3-Methylthreonine ODR 32 Table 6: Summary of Experimental versus Computational Results 33 Table 7: Proton Affinity of Cysteine and Serine 33 iv Acknowledgements I would to first thank Dr. J.C. Poutsma for his guidance and encouragement throughout my years in IonLab. I would also like to thank the members of IonLab for their support and collaboration. I would like to especially thank Izzy Owens, whose teamwork on this project was invaluable, as well as Zach Smith and Gwendyln Turner, for all their work in the computational aspects of the project. I would also like to thank Anwar Radwan for his continuous positive outlook and friendship throughout our time together in IonLab. I would like to thank both the College of William and Mary and the Charles Center for making this honors thesis possible. Lastly, I would like to thank the National Science Foundation for funding this project. v Chapter 1: Introduction 1.1 Acid Base Chemistry The acid-base properties of molecules are of interest to chemists for a variety of reasons. Thermodynamic properties such as acidity and basicity are partially responsible for governing the structure, bonding, and function of molecules. By determining these properties, we are able to get a clearer picture of the molecule’s overall structure and bonding behavior. These properties are useful in many different fields of study, ranging from medicine and biology to chemistry and nutrition.3-7 Acids and bases follow a typical reaction, which can be seen in (1), where a protonated conjugate acid (BH+) dissociates into a gaseous base (B) and a free proton (H+). !"#(%) → !(%) + "#; Δ" = -.; ∆0 = 0! (1) In Equation 1, proton affinity (PA) is the enthalpy change (Δ") for the reaction, and gas basicity (GB) is the Gibbs free energy (∆0) for the reaction. 8 This equation follows the Bronsted definition of acids and bases, where a base is a molecule that is capable of accepting a proton from a proton donor, while an acid is a molecule that is capable of donating a proton to a proton acceptor.9 Amino acids are of particular interest in acid-base chemistry study, as they are the building blocks of peptides and proteins. Although there are only twenty common protein amino acids that are incorporated into proteins in humans, there are many other non-protein amino acids that are of interest to study.10,11 Protein amino acids are those that are found in the main chains of peptides and proteins, while the non-protein amino acids are not found in protein main chains either because they do not have the specific transfer RNA and codon triplet needed, or because they do not have the correct post-translational modification.10 Studying these non-protein amino acids is important, as many of them have potential benefits in areas such as drug discovery, while others are toxic to 1 humans, secondary to their similar structures to protein amino acids.11 As these non-protein amino acids have similar structures to the common amino acids, it is of interest to see how these small differences in structure change the molecules bonding and acid-base properties. While acid-base reactions can be studied both in solution and in the gas-phase, gas-phase study is of interest due to the reactions solely depending on the acid-base strength of the reacting species, rather than on solvation effects.12 These reactions follow gas-phase acidity and gas-phase basicity scales, which are fundamentally different from the aqueous solution scale of pKa, due to solvation effects.12 On the gas-phase acidity/basicity scale, a stronger acid will have a smaller gas- phase acidity, and a stronger base will have a larger gas-phase basicity.12 Many of the experiments in gas-phase acid-base chemistry rely on the proton to evaluate the strength of the base. Acid-base reactions are based on the transfer of a proton and it is of interest to be able to measure the proton affinity of molecules to evaluate how strongly the proton will be attracted to a molecule of interest. This is discussed in detail in section 1.3, the kinetic method. 1.2 Previous Non-Protein Amino Acid Studies This lab has previously studied the thermochemical properties of various non-protein amino acids, including analogs of proline, lysine and arginine.5-7 All of these studies sought to determine how the thermochemical properties change with small structural changes to the amino acid.5-7 For proline, the study sought to determine the effects of increasing ring size on proton affinity.5 The proline analogs included L-azetidine-2-carboxylic acid, L-proline and L-pipecolic acid, which have 4-, 5-, and 6-membered ring nitrogen heterocycles, respectively.5 This study found that L-azetidine-2-carboylic acid, L-proline and L-pipecolic acid had proton affinities of 933 kJ/mol ± 6.3 kJ/mol, 941 kJ/mol ± 6.7 kJ/mol and 944 kJ/mol ± 6.7 kJ/mol, respectively.5 This 2 study was consistent with previous studies, which also found that increasing the size of a nitrogen heterocycle ring increased proton affinity.5,13,14 A second study by