Design and Application of MA’AT Analysis, a New Method for Conformational Analysis of Molecules in Solution

The molecular structures of biomolecules are highly associated with their cellular function. The determination of accurate structures is imperative to understanding how these molecules operate. However, methods to assign structures in solution have remained challenging for flexible molecules such as carbohydrates. This problem has led to a heavy reliance on theoretical calculations to produce models of solution conformations with limited or no experimental validation. This dissertation details the development and validation of a novel method, MA’AT analysis, to model conformational behavior in solution using a combination of NMR spin-couplings and density functional theory (DFT). The work presented here is separated into three distinct areas.

First, Chapter 2 details the development of the MA’AT method and its encoding into a user-friendly web-based application. MA’AT utilizes a set of parameterized equations that quantify the relationship between torsional angles or conformations with NMR spin-spin coupling constants (J-couplings) to produce population distributions. The theory and mathematical proofs underpinning the method are detailed in Chapter 2, along with a description of the testing, validation, and limitations of the method.

Conformational studies utilizing MA’AT are presented in Chapters 3–7. These studies developed new equations that describe the Karplus-like relationships between molecular conformation and coupling magnitudes as determined by DFT. Also discussed are the methods to obtain experimental J-couplings. The conformational models produced by MA’AT analysis are then compared to those obtained from molecular dynamic (MD) simulations and X-ray crystal structures.

Finally, Chapters 8–9 describe studies undertaken to validate the DFT-derived parameterized J-coupling equations used MA’AT analysis. Chapter 8 describes studies that utilized solid state 13C NMR (ssNMR) to measure JCC spin-coupling constants in molecules having essentially fixed conformations. In these cases, experimentally-measured J-couplings were directly compared to those calculated from DFT. Finally, Chapter 9 describes a computational study on the torsional dependence of non-conventional coupling pathways for O-glycosidic linkages.