Theses and Dissertations from UMD
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Item Catechols as Membrane Anion Transporters(2009) Berezin, Sofya; Davis, Jeffery T.; Chemistry; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)ABSTRACT Title of Document: CATECHOLS AS MEMBRANE ANION TRANSPORTERS Sofya Berezin, Doctor of Philosophy, 2009 Directed By: Professor, Jeffery T. Davis, Department of Chemistry and Biochemistry Synthetic anion transporters have potential as antimicrobials, extractants, sensors, etc. Anionophores may also help us understand how natural systems move ions across hydrophobic barriers. While bacterial siderophores and synthetic analogues use catecholates for Fe3+ uptake, this work reports of catechols facilitating biomembrane transport of anions. We demonstrate that simple bis-catechol III-25 is an anion transporter whose activity depends on catechol's substitution and amphiphilicity. We also describe liposomal assays and devised quantitative description that allows one to study facilitated anion transport. These assays indicate that selectivity of III-25 follows the Hofmeister bias: anions which are easier to dehydrate are made more permeable to the membrane by this bis-catechol. We believe that our description of the ion selectivity and mechanism for III-25 opens an outstanding opportunity for those interested in determining the selectivity and mechanism for other synthetic and natural biomembrane ion transporters. In the beginning of this project we investigated number of simple amides and phenols to evaluate their relative affinity and stoichiometry of interaction with Cl- anion. ESI-MS and 1H NMR analysis showed that a dimer, catechol2*Cl-, was the major complex formed when TBA+Cl- was mixed with excess catechol. Based on this finding we attached two catechols to a TREN scaffold. A hydrophobic alkyl amide groups were linked to TREN's third position. Surprisingly, this simple design led to the active analogs III-23 - III-26. A medium-length, III-25, was the most active compound, indicating that ion transport ability depends on the ability to partition into the biomembrane. Finally, we noticed that the experimentally observed weak dependence of the transport rates on the anion's hydration energy, namely, kAnion decreasing in the order ClO4- > I- > NO3- > Br- > Cl-, is also seen for some of Nature's anion transporters. Thus, anion permeation into the CFTR chloride channel shows a similar trend. We also observed a nonlinear dependence of kAnion on the concentration of bis-catechol. These findings led us to believe that self-association of III-25 provides transient pores that allow permeation without requiring complete dehydration of the inorganic anions. Future efforts will include incorporating selectivity filters into these bis-catechols to help overcome the Hofmeister bias.Item USING "SMALL" MOLECULES AS TRANSMEMBRANE ANION TRANSPORTERS(2009) Okunola, Oluyomi Adeola; Davis, Jeffery T; Chemistry; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)Functional, "small" molecule anion transporters have been identified and developed from natural products and synthetic organic compounds. The major discoveries include the design of a transmembrane Cl- transporter whose activity is pH-tunable, a NO3- transporter that displays unique selectivity for NO3- over Cl- anions, and a series of small molecules that efficiently transport HCO3- across liposomal membranes via a HCO3-/Cl- exchange mechanism. An assay for detecting transmembrane HCO3- transport using paramagnetic Mn2+ and 13C NMR is also described. Modulated Cl- transport was achieved by lipophilic calix[4]arene amides 2.2-2.4, all in the cone conformation. Modulation was achieved through functional group modification to one of the four side-chains. The cone conformation was confirmed by both 1H NMR and X-ray crystallography. Significantly, Cl- transport was gated by pH in the presence of triamido calixarene TAC-OH 2.3, which possesses a phenolic hydroxyl group. Using fluorescence assays, the rate of Cl- transport by TAC-OH 2.3 across liposomal membranes decreased with increasing pH, while transport rate by cone-H 2.2a, lacking an OH group, was not affected by pH. Nitrate was selectively transported over Cl- in the presence of nitro tripod 3.1, a small molecule receptor for both anions. The selective transport of NO3- by 3.1 is a significant discovery as most known synthetic Cl- transporters also transport NO3- ions and vice versa. Nitrate transport across liposomal membranes was confirmed by enzyme-coupled and fluorescence assays. Tripod 3.1 induced an increase in the intravesicular pH of liposomes that were not experiencing a pH gradient, while no pH changes occurred in the presence of calixarene 2.1 a known Cl- and NO3- transporter. This result suggests that 3.1 is an H+/NO3- symporter. Transmembrane HCO3- transport was achieved using the natural product, prodigiosin, 4.1, and synthetic isophthalamides 4.2-4.4. The Cl-/HCO3- exchange mechanism by which compounds 4.1-4.4 transport HCO3- was elucidated by ISE and NMR assays. The 13C NMR assay provided direct evidence for HCO3- transport in the presence of paramagnetic Mn2+ ions, and was adaptable to various assay conditions.