The atomic resolution structure of a low populated and transiently formed on-pathway folding intermediate of the FF domain from human HYPA/FBP11 has recently been reported[1]. The structure was determined on the basis of backbone chemical shift and bond vector orientation restraints measured on the ‘invisible’ intermediate state using relaxation dispersion nuclear magnetic resonance (NMR) spectroscopy that were subsequently input into the data-base structure determination program CS-Rosetta. This thesis focuses on the cross-validation of the structure so produced. We present here the solution NMR structure of a mimic of the folding intermediate that is highly populated in solution, obtained from the wild-type domain by protein mutagenesis. The ensemble of structures generated of the mimic are within 2Å of the relaxation dispersion/CS-Rosetta structures of the intermediate, with the non-native interactions in the intermediate also observed in the mimic. The results presented in this thesis strongly confirm the structure of the FF domain folding intermediate, in particular, and validate the use of relaxation dispersion derived restraints in structural studies of invisible excited states, in general.

The number of gene products available for structural and functional study is increasing at an unprecedented rate as a result of the successful whole genome sequencing projects. Systematic structure determination of proteins on a genomic scale, called structural genomics, can significantly contribute to the field of protein science and to functional annotation of newly identified genes. This thesis covers different aspects of protein production in Eschericiha coli for structural studies in the context of structural genomics. Protocols have been downscaled and standardized to allow for a rapid assessment of the production characteristics for multiple proteins in parallel under a number of different conditions. Foremost, the ability of different proteins and peptide tags to affect the solubility of the recombinant protein when produced as fusion proteins has been systematically studied. Large differences in the success-rate for production of soluble protein in E. coli were found depending on the fusion partner used, with a more than two-fold increase in the number of proteins produced as soluble when comparing the best and the poorest fusion tags. For different constructs with a histidine tag, commonly used to facilitate protein purification, large differences in yield depending on the design of the expression vector were found. When comparing different fusion proteins produced from identical expression vectors, fusions to the GB1 domain were found to result in the highest yield of purified target protein, on average 25 % higher than any of the other fusions. The suitability for further structural studies was tested at an intermediate scale for proteins that were identified as soluble in the expression screening. For this purpose, protocols for rapid purification and biophysical characterization using nuclear magnetic resonance and circular dichroism spectroscopy were developed and tested on 19 proteins, of which four were structured. / QC 20100826

Thesis (M. Sc.)--Carleton University, 2004. / Includes bibliographical references (p. 130-140). Also available in electronic format on the Internet.

Thesis (Ph.D. (Chemistry)) - Rhodes University, 2008.

Thesis (Ph.D.) -- University of Western Sydney, 2008. / A thesis submitted to the University of Western Sydney, College of Health and Science, School of Natural Sciences, in fulfilment of the requirements for the degree of Doctor of Philosophy. Includes bibliographical references.

Thesis (M.Sc.). / Written for the Dept. of Chemistry. Title from title page of PDF (viewed 2009/06/23). Includes bibliographical references.

Thesis (Ph. D.) -- University of Texas Southwestern Medical Center at Dallas, 2005. / Vita. Bibliography: 209-224.

Thesis (Ph. D.)--University of Wisconsin--Madison, 1983. / Typescript. "January 1983"--Cover of photocopy. Two p. of original are reproduced on 1 p. of photocopy. eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references (leaves 151-152).

Thesis (Ph. D.)--Ohio State University, 2006. / Title from first page of PDF file. Includes bibliographical references (p. 355-382).

Thesis (Msc Food Sc (Food Science))--University of Stellenbosch, 2005. / Near infrared (NIR) spectroscopy prediction models were developed for the rapid determination of the peroxide value, free fatty acid content, specific extinction coefficients at 232 and 270 nm, thiobarbituric acid reactive substances value, pigment, total polyphenol, oleic acid, linoleic acid and saturated fatty acid contents ...