No description available.

No description available.

No description available.

No description available.

This work is a part of a large project in our laboratory that is aimed toward characterization of prokaryotic transcription regulators from different families and their interactions with small-molecule effectors. My study was focused of IclR family of transcriprion regulators, specifically on its founding member Isocytrate Lyase Regulator (IclR) from E.coli and AllR regulator from E.coli, which share 42% sequence identity with IclR. I used a combination of biophysical, biochemical and structural biology techniques to explore the mechanisms by which IclR and AllR interact with their effectors. I performed site-directed mutagenesis experiments in order to research the role of individual amino acids in interaction of AllR regulator with its previously identified effector glyoxylate and to test whether oligomerization plays a role in effector-induced signal transduction by AllR. Using differential light scattering, which allows high-throughput screening of small molecules for thermostabilization of proteins, I identified potential effctors for the IclR regulator. The physiological relevance of these candidate molecules was tested in-vitro and in-vivo and their interaction with IclR was characterized by Isothermal Titration Calorimetry and X-ray Crystallography.

This work is a part of a large project in our laboratory that is aimed toward characterization of prokaryotic transcription regulators from different families and their interactions with small-molecule effectors. My study was focused of IclR family of transcriprion regulators, specifically on its founding member Isocytrate Lyase Regulator (IclR) from E.coli and AllR regulator from E.coli, which share 42% sequence identity with IclR. I used a combination of biophysical, biochemical and structural biology techniques to explore the mechanisms by which IclR and AllR interact with their effectors. I performed site-directed mutagenesis experiments in order to research the role of individual amino acids in interaction of AllR regulator with its previously identified effector glyoxylate and to test whether oligomerization plays a role in effector-induced signal transduction by AllR. Using differential light scattering, which allows high-throughput screening of small molecules for thermostabilization of proteins, I identified potential effctors for the IclR regulator. The physiological relevance of these candidate molecules was tested in-vitro and in-vivo and their interaction with IclR was characterized by Isothermal Titration Calorimetry and X-ray Crystallography.

The biological significance of macrocyclic complexes has been recognized since they were first synthesized by Neil Curtis. They have the potential to play a critical role in mimicking metalloprotein active sites. Nine Curtis macrocyclic complexes have been studied using X-ray crystallographic techniques. Their structures have been solved and comparisons of the results have been made. Biological importance is also true of the macrocyclic counterpart; side-off and end-off compartmental ligands. In some circumstances these types of ligands are more appropriate because they have extra flexibility due to their pendant arms not being fixed in place by another head-unit, like a traditional macrocycle. The synthesis of a proposed compartmental ligand; 2,2-(N,Nʼ-bis(benzimidazole-2-ylmethyl)methylamine-5,5ʼ-di-tert-butyl-3,3ʼmethanediyl-dibenzyl alcohol (Ligand 1(L1)), has been proposed and outlined. The pendant arms: bis(benzimidazole-2-ylmethyl)amine (BBIM), were successfully synthesized and characterized with 1H NMR, IR and X-ray crystallography. The head-unit: 5,5ʼ-Di-tert-butyl-2,2ʼ-dihydroxy-3,3ʼ-methanediyl-dibenzene methanol (DHTMBA), of L1 was synthesized and characterized using 1H NMR, IR and mass spectrometry. A similar head-unit; 5,5ʼ-Di-methyl-2,2ʼ-dihydroxy-3,3ʼ-methanediyl-dibenzene methanol (DHMMBA), was synthesized in an effort to shorten the synthetic time of the head-unit. This was consequently converted to the chlorine analogue; 3,3ʼ-Bis(chloromethyl)-5,5ʼ-dimethyl-2,2ʼ-methane-diyldiphenol (Cl-DHMMB), and characterized with 1H NMR, IR and X-ray crystallography. Efforts were made to synthesize Ligand 1, but due to synthetic difficulties and time restraints this proved unsuccessful. Suggestions have been made to develop this synthesis.

Thesis (Ph. D.)--University of Washington. / Bibliography: l. 50-52, 192-195.

Thesis (Ph.D.) -- McMaster University, 1998. / Includes bibliographical references (p. 134-135). Also available via World Wide Web.

Thesis (Ph.D.) - University of St Andrews, November 2006.