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Intermolecular and Intramolecular Stable Isotope Studies in Alanine


Dallas, Brooke Hillary (2024) Intermolecular and Intramolecular Stable Isotope Studies in Alanine. Dissertation (Ph.D.), California Institute of Technology. doi:10.7907/xqdx-pp86.


In Chapter 1 of this thesis, we give an introduction to this body of work, providing some background for context.

In Chapter 2, we present a set of theoretical predictions for the carbon isotope distribution between equilibrated carbon sites of alanine and pyruvate. We start with the simplest possible theoretical treatment, and work progressively through higher levels of theory, showing consistency in the direction and magnitude of expected fractionation across these treatments.

In Chapter 3, we present our experimental work to confirm the predictions made in Chapter 2 by measuring the δ13C of the α carbon site in alanine that has undergone equilibration with the analogous carbon site in pyruvate via the alanine transaminase enzyme (ALT).

In Chapter 4, we describe the process that led to our (re)discovery of β-hydrogen-deuterium exchange in amino acids catalyzed by transaminases. We then provide a literature review on the small body of historical work on this system, which took place primarily during the 1960s and 70s. This literature summary provides the background necessary for the reader to appreciate our experimental work presented in the next chapter.

In Chapter 5, we present novel 1H NMR and 13C NMR experimental observations of intermolecular hydrogen isotope exchange between water and the α and β carbon sites of alanine, as well as intramolecular hydrogen isotope exchange between the α and β carbons, all of which is catalyzed by alanine transaminase (ALT). These experiments track the abundances of eight isotopically distinct alanine species varying in their position and/or number of hydrogen isotopes over a series of reactions differing in initial alanine isotopic composition and initial water isotopic composition. With the data collected we are able to determine up to thirteen rate constants and ten equilibrium constants describing the transfer of hydrogen and deuterium amongst these eight isotopic variants and water, as well as the thermodynamic equilibrium constants between them.

Item Type:Thesis (Dissertation (Ph.D.))
Subject Keywords:stable isotope; geochemistry; carbon; hydrogen; deuterium; enzyme kinetics; isotope exchange; density functional theory; electronic structure; nwchem; enzymology; biochemistry; amino acid; alanine; pyruvate; metabolism; citric acid cycle; transamination; transaminase; biogeochemistry; organic geochemistry; position specific isotope analysis; mass spectrometry; nuclear magnetic resonance; nmr
Degree Grantor:California Institute of Technology
Division:Geological and Planetary Sciences
Major Option:Geology
Thesis Availability:Public (worldwide access)
Research Advisor(s):
  • Eiler, John M.
Thesis Committee:
  • Sessions, Alex L. (chair)
  • Blake, Geoffrey A.
  • Leadbetter, Jared R.
  • Eiler, John M.
Defense Date:8 May 2024
Non-Caltech Author Email:brookehdallas (AT)
Funding AgencyGrant Number
Department of Energy (DOE)DE-SC0016561
Record Number:CaltechTHESIS:05252024-201300283
Persistent URL:
Related URLs:
URLURL TypeDescription Material adapted for Chapter 2
Dallas, Brooke Hillary0000-0002-1313-3270
Default Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:16426
Deposited By: Brooke Dallas
Deposited On:30 May 2024 00:06
Last Modified:12 Jun 2024 21:42

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