Thesis (Master)--İzmir Institute of Technology, İzmir, 2004. / Includes bibliographical references (leaves. 70-73).

Soluble guanylyl cyclase (sGC), the nitric oxide (NO) receptor, is a 150 kDa heterodimeric multi-domain protein that contains heme in the β subunit. Binding of NO to heme leads to rupture of the proximal histidine bond, increased catalytic conversion of GTP to cGMP at a distant guanylyl cyclase catalytic domain, and vasodilation through cGMP signaling. The structure of sGC has not been determined, and little is known about the mechanism by which NO binding to heme leads to increased catalysis. The small molecule YC-1 is known to stimulate sGC activity, but the exact YC-1 binding site and mechanism of action are unknown. Using truncated constructs of Manduca sexta (Ms) sGC lacking the catalytic domain, conformational changes upon YC-1 and NO-binding were characterized using analytical ultracentrifugation and small-angle X-ray scattering. Chemical cross-linking and high-resolution mass spectrometry was used to obtain distance restraints which, when combined with homology models, have provided the first model of sGC domain arrangement and revealed important information about domain-domain interactions. Truncated Ms sGC is highly elongated, contains a coiled-coil in a parallel arrangement, and contains a direct interface between the β H-NOX (Heme Nitric oxide/Oxygen binding domain) and the coiled-coil, and between the β H-NOX and α PAS (Per-arnt-sim) domains. Experiments using analytical ultracentrifugation, fluorescence anisotropy and native mass spectrometry have revealed the YC-1 binding site to be located within the α PAS domain. Additionally, measurement of the kinetics of heme loss and the heme reduction potential were performed to investigate the instability of oxidized sGC heme.

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A set of water-soluble 4,4-difluoro-4-bora-3a,4a-diaza-s-indacene (BODIPY) derivatives, has been prepared and their spectroscopic properties examined. These dyes can be used as either donor or acceptor in synthesis of through-bond energy transfer cassettes. Sulfonation conditions were developed for several BODIPY dyes to give the mono-sulfonated and di-sulfonated products. Compounds with an aryl iodide could be used for organometallic couplings. Similarly, BODIPYs with an aromatic bromide, but also two chlorine atoms could be replaced via SNAr reactions. The amine sulfonated BODIPY is amenable to couple to biomolecules via acylation reactions. A diazotization/azide reaction sequence was used to convert the amines into azides; the latter may be functionalized via click reactions. Spectral data for these materials indicates they are highly fluorescent probes in aqueous environments. We have also prepared some lipophilic BODIPY derivatives, which can be used for SNAr reactions and make some through-bond, energy transfer cassettes. DichloroBODIPYs can also be used for labeling proteins successfully.

Phase selective soluble polymers are useful in organic synthesis because they simplify purification and separation processes. Such selective soluble polymers enable the use of Green chemistry principles to be utilized as ways to simplify catalyst, reagent, and product recovery. Polyisobutylene oligomers serve as examples of such polymers. Vinyl terminated polyisobutylene (PIB) oligomers can be easily transformed into a variety of end-functionalized PIB oligomers. Previous work has shown that PIB oligomers possess nonpolar phase selective solubility that allows them to be used as polymer supports for ligands and catalysts in liquid/liquid biphasic systems. This dissertation focuses on the use of PIB oligomers as supports for a salen Cr(III) complex, a Hoveyda-Grubbs 2nd generation catalyst, and a N-heterocyclic carbene. The syntheses of these PIB-supported ligands and catalysts are simple and straightforward. The synthetic products and the intermediates in these syntheses can all be readily analyzed and monitored by conventional spectroscopic methods. The activity of the PIBsupported catalysts is shown to be analogous to that of other soluble polymer supported catalysts or their non-supported analogs. The PIB-bound catalysts can be separated from products by a latent biphasic, liquid/liquid extractions, or product self-separation systems. The recovered PIB-bound catalysts can then be recycled multiple times.