<p>Medical advances often come as a result of understanding the underlying mechanisms of life. Life, in this sense, happens at various scales. A very complex and interesting one is the molecular scale. Understanding life’s mechanistic details at this level will provide the most promising therapies to modern ailments. Because of structure and function’s close relationship, knowledge of macromolecular structure provides invaluable insight into molecular mechanism.</p><p>A major tool used to get structural information at the molecular scale is X-ray crystallography. Such experiments result in an electron density map from which a model is built. Building such a model is a difficult task, especially at low resolu- tion where detailed features in the electron density deteriorate making it difficult to interpret. However, many advances in the field have greatly eased the model build- ing task, in fact, at high resolutions it has become automated. However, human inspection is still required to get a correct solution.</p><p>The largest boon to model building has been the application of structural knowl- edge. A prominent example is bond and dihedral angles. We often know what is absolutely not allowed and often convince ourselves we know everything that is al- lowed. This work focuses on the fuzzy border between allowed and disallowed. The hypothesis is that rare structural conformations exist but one needs to take great care in modeling them.</p><p>This work has two major components – rotamers (protein sidechain conformation)</p><p>and Hoogsteen base pairing in DNA. I first describe methods used to gain empirical knowledge about rotamers and how that knowledge is used in model validation. Part of this knowledge is rotamer-dependent bond angle deviations. I describe how the observation and quantitation of these deviations is used in a novel set of restraints in protein structure refinement. To provide structural context to rare rotamers, I describe where and why some occur.</p><p>My DNA work has focused on Hoogsteen base pairing. I describe a collaborative survey of existing Hoogsteen base pairs in the PDB. Lessons learned during the survey led to the other DNA topic, the detection and correction of mismodeled purines. I identified Hoogsteens in the PDB mismodeled as Watson-Crick base pairs. This work underscores that Hoogsteens are extremely rare but nonetheless do occur.</p><p>The fuzzy borderland between allowed and disallowed is a strange place filled with the most interesting structural features. My work here has focused on this area, bringing into view many rare conformations. Going forward we need to ensure that conformational frequency is taken into account during model building, refinement, and validation.</p> / Dissertation
| Identifer | oai:union.ndltd.org:DUKE/oai:dukespace.lib.duke.edu:10161/11345 |
| Date | January 2015 |
| Creators | Hintze, Bradley Joel |
| Contributors | Richardson, Jane S, Richardson, David C |
| Source Sets | Duke University |
| Detected Language | English |
| Type | Dissertation |
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