User:Szeichner/Position-specific isotope analysis


 * my contributions are in bold, to distinguish from Elliott's contributions

Proposal:
''We want to flesh out the page for position specific isotope analysis. There is some information already on the wikipedia page, but much to add about motivation (e.g., how position specific isotopes can reveal mechanism), instrumentation (GC, HPLC, Orbitrap, NMR), and applications (e.g., Caj’s methionine paper). We plan to elaborate on the theory behind site specific enrichments, connecting this to biological and abiological mechanisms of fractionation that can only be revealed by site specific measurements. This will then be coupled to the aforementioned instrumentation section. The final piece of the site will be the applications and case studies, potentially a closer look at photosynthetic site specific anomalies, amino acids (Abelson&Hoering and Caj’s paper), and fatty acids (Melzer & Schmidt). As this is a rich topic, we propose to work on this larger endeavor together and split up the work. I will focus my section on the examples and applications of this work; Elliott will focus on the theory and instrumentation (although we may iterate on this division of labor as we progress through the process).''

Introduction:

 * Stable isotope analysis (bulk versus compound versus position specific)
 * Figure here tracing heavy isotopes
 * Applications/utility:
 * Isotope labeling studies (e..g, doping)
 * Biomolecule origins

Theory

 * Site specificity can deviate from stochastic distribution, and these deviations are meaningful.
 * They are controlled by the equilibria, branch points, and isotope effects along chemical reaction networks.
 * Lighter isotopes are generally preferred kinetically but heavier isotopes are preferred in higher energy bonding environments. These preferences are atom-specific.


 * Biological fractionation
 * Enzymes show a preference for lighter isotopes based on their faster rate constants. These are related to the energy state of TS (link to some other wiki page?)
 * Intrinsic effects vs. expressed effects
 * How can we measure these?
 * Direct comparison: rate of monoisotopic vs. singly substituted
 * Internal competition: measure isotope ratios over reaction progress
 * Equilibrium perturbation: measuring equilibrium isotope effects inside enzymes


 * Abiological fractionation
 * Thermogenic isotope effects
 * FIscher Trospch-type synthesis, Strecker synthesis

Section 1: On-line Measurements

 * Chromatography
 * The huge FTMS at the national lab in N. California (forget its name)
 * Orbitrap
 * NMR
 * Pyrolysis-GC-C-IRMS
 * Pyrolysis-GC-C-IRMS

Section 2: Off-line Measurements

 * Before the advent of site-specific measurements on whole molecules, we needed to first break it down by chemical degradation.
 * Chemical degradation:
 * Ozonolysis (Monson & Hayes)
 * Pyrolysis (Pyruvate and acetate)
 * Enzymatic degradation:
 * glucose-6-phosphate, pyruvate decarboxylase, etc. (any Cleland paper).

Case studies

 * Photosynthesis
 * Phosphoenolpyruvate carboxylase (PEPC) as a good example of EIEs and normal and inverse KIEs
 * Start with a simple example of equilibrium (CO2 → HCO3-) prefers the heavier isotope because of molecular geometry.
 * Shift to this heavy carbon incorporating into aspartic acid via PEPC (O’Leary, 1981), we see this in site-specific measurements
 * When the enzyme uses bicarbonate it actually expresses a normal isotope effect  on the bicarbonate but an inverse effect on the C1 of PEP.
 * This shows how isotope effects really are atom-specific

Transition: We see that intramolecular patterns arise in most AA’s (Abelson & Hoering):

(Do wiki articles need transitions at all?)


 * Amino acids
 * Carboxylic acid enrichment (Abelson & Hoering)
 * Methionine and its sources (Neubauer et al)


 * Fatty acids
 * Melzer and Schmidt