Thesis (Ph. D.)--Massachusetts Institute of Technology, Dept. of Chemistry, 2007. / Vita. / Includes bibliographical references. / Two distinct structural scaffolds elaborated from Schiff-base macrocycles were designed to study the proton-coupled electron transfer chemistry of 0-0 bond forming and activation chemistry. The "Hangman" architecture is composed of hydrogen-bonding functionalities poised over a redox active manganese salophen or salen platform. The complexes proved to be proficient catalase mimics (disproportionation of hydrogen peroxide to water and oxygen). Detailed spectroscopic, computational, and structure-function relationship studies elucidated the key redox, steric, and secondary coordination sphere effects for optimal catalytic ability. The incorporation a chiral backbone into the macrocycle led to catalysts that perform enantioselective epoxidation of unfunctionalized olefins. A macrocycle with an amide, imine, and bisphenolic functionalities was also incorporated as the redox platform in the Hangman framework; the manganese complex also performed catalytic oxygen atom transfer to olefins. The second framework, dubbed "Pacman", is composed of two salen platforms linked cofacially by rigid pillars xanthene or dibenzofuran. A series of bimetallic complexes, including chromium, iron, manganese, cobalt, copper, and zinc were generated. Mossbauer spectroscopy was used in the characterization of the iron salen complexes, which were also examined for photolytic oxidation chemistry. / by Jenny Yue-fon Yang. / Ph.D.

Thesis (M.S.)--Massachusetts Institute of Technology, Dept. of Chemistry, 1997. / Includes bibliographical references. / by John Liamanto. / M.S.

Thesis (Ph. D.)--Massachusetts Institute of Technology, Dept. of Chemistry, 2006. / Vita. / Includes bibliographical references. / The biosynthesis of asparagine-linked glycoproteins, highly conserved throughout all eukaryotes, requires a dolichylpyrophosphate-linked tetradecasaccharide precursor (Dol-PP-GlcNAc2Man9Glc3), from which the tetradecasaccharide is transferred co-translationally to nascent polypeptides in the lumen of the ER by the multimeric membrane-bound enzyme, oligosaccharyl transferase (OT). The saccharide donor is assembled by a series of membrane-bound enzymes, which together comprise the dolichol pathway. Despite over two decades of genetic and bioinformatics approaches that have identified the vast majority of dolichol pathway genes in yeast, the roles of two mannosyltransferases in the pathway, Alg2 and Algl 1, remained ambiguous. This thesis describes the biochemical studies that were carried out to clarify these roles. The substrate specificity of Algl, the first mannosyltransferase in the pathway, was studied, and this enzyme was also used as a tool to prepare Man1 ,4-GlcNAc2-PP-Dol from synthetic GlcNAc2-PP-Dol. Access to this trisaccharide intermediate facilitated the characterization of Alg2 function, proposed to be involved in addition of the second and/or third mannose. / (cont.) A cell membrane fraction isolated from E. coli overexpressing thioredoxin-tagged Alg2 was used to demonstrate that this enzyme carries out an al 1,3-mannosylation, followed by an al,6-mannosylation, to form the branched pentasaccharide intermediate of the dolichol pathway. Having the means to access this intermediate chemoenzymatically, it was thus possible to define the function of Algl 1, which had similarly been proposed to catalyze addition of the fourth and/or fifth mannose. Using the same procedure, TRX-Algll 1 was shown to catalyze two sequential al 1,2-mannosylations onto the a 1,3-branch of the pentasaccharide substrate to afford the heptasaccharide intermediate. The elucidation of the dual function of each of these enzymes thus completes the identification of the entire ensemble of glycosyltransferases that comprise the dolichol pathway. Finally, peptidyl mimics based on the consensus site for glycosylation by OT on nascent polypeptides, Asn-Xaa-Thr/Ser, were designed and evaluated. / (cont.) Both substrate-based peptide isosteres and product-based neoglycoconjugates were used to investigate the conformational and stereoelectronic preferences of OT binding. Neoglycoconjugates also showed promise as inhibitors of the deglycosylating enzyme Peptide: N-glycanase (PNGase), which aids in the degradation of misfolded proteins in the secretory pathway. / by Mary K. O'Reilly. / Ph.D.

Thesis (Ph. D.)--Massachusetts Institute of Technology, Dept. of Chemistry, 2009. / Vita. Cataloged from PDF version of thesis. / Includes bibliographical references. / Chapter 1. An Introduction to Fluorescent Zn(II) Sensors and Their Applications in Biological Systems This chapter opens with an overview of the numerous roles of zinc in biology, with an emphasis on labile Zn(II), that is, loosely bound or kinetically available stores of the ion. The synthesis, characterization, and application of fluorescent sensors for Zn(II) detection constitute an active field of research. A survey of the history and development of these sensors is presented, along with a sample of the knowledge acquired through their use in biological systems. An examination of methods for the subcellular localization of fluorescent probes is included. Chapter 2. Solution and Fluorescence Properties of Symmetric Dipicolylamine- Containing Dichlorofluorescein-Based Zn2+ Sensors The syntheses of five new analogues of the 2',7'-dichlorofluorescein-based Zn2+ sensor Zinpyr-1 (ZP1) are reported. Analysis of these fluorophores by absorption and emission spectroscopy, in combination with potentiometric titrations, produced a detailed molecular picture of the Zn2+ and H+ binding properties of the ZP1 family of sensors. The two separate N30 donor atom sets on ZP1 converge to form binding pockets in which all four heteroatoms participate in coordination to either Zn2+ or protons. The position of the pyridyl group nitrogen atom, 2-pyridyl or 4-pyridyl, has a large impact on the fluorescence response of the dyes to protons in spite of relatively small changes in pKa values. Despite the structural complexity of ZP1, we provide evidence that the pyridyl arms of the DPA appendages participate in the quenching process, in addition to the contribution from the tertiary nitrogen amine atom. / (cont.) Potentiometric titrations reveal ZP1 dissociation constants (Kd) for Zn2 + of 0.04 pM and 1.2 nM for binding to the first and second binding pockets of the ligand, respectively, the second of which correlates with the value observed by fluorescence titration. Chapter 3. Subtle Modification of 2,2-Dipicolylamine Lowers the Affinity and Improves the Turn-On of Zn(II)-Selective Fluorescent Sensors The spectroscopic and proton- and Zn(II)-binding properties of two new members of the Zinpyr family of fluorescent sensors are reported. In ZP1B and ZP3B, a (2-picolyl)(4- picolyl)amine (2,4-DPA) moiety is installed in place of the di(2-picolyl)amine (2,2-DPA) ligand used in the parent sensors ZP1 and ZP3. This modification has the benefit of both lowering the proton-induced turn-on at physiological pH levels and altering the Zn(II) affinity so as to detect only the most concentrated stores of this ion in biological samples. Comparison of the proton affinities of all four sensors, as determined by potentiometric titrations, contributes to our understanding of the solution properties of this family of sensors. 5 Chapter 4. Labeling of the Cell Membrane with Fluorescent Sensors to Track Extracellular Zn(II) With the goal of labeling the plasma membrane of living cells to track fluctuations in extracellular Zn(II), the sulfosuccinimidyl ester of Zinpyr-1, ZPlss, was synthesized. Incubation of HeLa cells with this probe does result in fluorescent labeling of the cell surface but no response to exogenous Zn(II) was observed. Partial syntheses of potentially more sensitive versions of ZP Iss are also reported. / (cont.) Appendix 1. Synthesis of Components for the Construction of Tethered Zn(II)- Responsive Fluorophores This appendix describes efforts to synthesize membrane-targeted fluorescent Zn(II) sensors based on two different coupling partners for azide-bearing cellular components. Zinpyr derivatives with a water-soluble amine tethering group were synthesized as precursors to the final phosphine and strained cyclooctyne constructs. The reported compounds should serve as useful synthons for future studies in probe localization. Appendix 2. Miscellaneous NMR Spectra / by Brian Alexander Wong. / Ph.D.

Thesis (Ph.D.)--Massachusetts Institute of Technology, Dept. of Chemistry, 2000. / Includes bibliographical references. / Semiconductor quantum dots or nanocrystals have size dependent optical and electronic properties that arise from quantum confinement. While the quantum size effect is reasonably well understood, the effect of abrupt interface between the nanocrystal and its dielectric environment is not. In this thesis we study how the dielectric environment affects the quantum dot electronic structure, the optical absorption ~ross-section, charge separation, and transport in cadmium selenide colloidal quantum dots. The electronic states and optical absorption cross-section are found to be less sensitive to changes in the dielectric environment than predicted from theory unless screening from the ligand shell is taken into account. The absolute absorption cross section is measured as a function of quantum dot size; excellent agreement with theory is obtained for absorption far above the band edge. Three-dimensional close packed solids of quantum dots are predicted to act as model artificial solids. Optical absorption measurements indicate that the electronic states of CdSe quantum dots separated by 11 angstroms or more are essentially uncoupled. Photoconductivity measurements suggest that photoexcited quantum confined excitons are ionized by the applied field with a rate that depends on both the size and surface passivation of the quantum dots. The charge generation efficiency decreases with increasing temperature as non-radiative and radiative recombination pathways increasingly compete with charge separation. A simple tunneling model for the initial charge separation step is presented that qualitatively reproduces both the size and surface dependence of the photoconductivity as a function of applied electric field. Finally, we report observations of amplified spontaneous emission from quantum dot solids. The stimulated emission is tunable with quantum dot size and does not sensitively depend upon surface passivation. These measurements demonstrate the feasibility of nanocrystal quantum dot lasers and amplifiers. / by Catherine A. Leatherdale. / Ph.D.

Thesis (Ph. D.)--Massachusetts Institute of Technology, Dept. of Chemistry, 1996. / by Todd Sterling Anderson. / Ph.D.

Thesis (Ph. D.)--Massachusetts Institute of Technology, Dept. of Chemistry, 1996. / Includes bibliographical references (p. 82-89). / by Yoko Kato. / Ph.D.

Thesis (Ph. D.)--Massachusetts Institute of Technology, Dept. of Chemistry, 2008. / Vita. / Includes bibliographical references. / Nitric oxide (NO) is a molecule that is essential for life and regulates both beneficial and harmful processes. Because this gaseous radical influences many aspects of health and disease, we wish to explore the relationship between NO and physiology/pathophysiology. To this end, we seek to create tools for the fluorescent imaging of NO in vivo. We have adapted an existing small molecule-based sensor for more biologically relevant applications by including it within a polymeric film. We have also developed turn-on fluorescent sensors for NO based on conjugated polymers, which demonstrated good selectivity and sensitivity for this analyte. In addition, we have prepared a related sensor that will detect nitroxyl (HNO) but not NO. These systems demonstrate the versatility and value of the conjugated polymer scaffold for sensing applications. Many targets of the diatomic radical NO contain redox active units, such as transition metals or thiolate ligands. To gain insight into how NO might regulate biological processes by interacting with these redox active species, we have initiated a fundamental study of the reactivity of NO with transition metal thiolate model complexes. Our explorations in this field have yielded unique nickel and cobalt nitrosyl species with atypical electronic and structural parameters. These studies have suggested intermediates for the more biologically relevant iron nitrosyl complexes that have not yet been observed may exist. Furthermore, the NO chemistry of these small molecule nickel and cobalt thiolate complexes may guide future biological investigations into the regulation of nickel and cobalt metalloproteins by NO. / Andrew Gregory Tennyson. / Ph.D.

Thesis (Ph. D.)--Massachusetts Institute of Technology, Dept. of Chemistry, 2007. / This electronic version was submitted by the student author. The certified thesis is available in the Institute Archives and Special Collections. / MIT Institute Archives copy includes accompanying CDROM with copy of thesis in .pdf format. / "September 2007." Vita. / Includes bibliographical references. / Non-heme carboxylate-bridged diiron centers in the hydroxylase components of the bacterial multicomponent monooxygenases activate dioxygen at structurally homologous active sites. Catalysis requires the management of four substrates: electrons, protons, dioxygen, and hydrocarbons. Protein component complexes control the delivery of these substrates to the diiron center in the hydroxylase ensuring selective hydrocarbon oxidation. A detailed mechanistic understanding of structural and chemical consequences of such interactions is a significant challenge. This thesis begins with an overview of our current understanding of these processes. The discussion is primarily on the methane monooxygenase systems (MMO) because these have been the most extensively studied BMMs to date. Recent results for the toluene/o-xylene monooxygenase (ToMO) and phenol hydroxylase systems from Pseudomonas sporium OX1 are also briefly summarized, the former being the research focus of this dissertation. Restricting access to the diiron center in ToMOH and other non-heme carboxylate-bridged diiron proteins was proposed to facilitate observation of oxygenated intermediates. To examine this hypothesis, dioxygen activation in ToMOH mutants that were predicted to occlude this channel was investigated by rapid-freeze quench (RFQ) EPR, Mossbauer, and ENDOR spectroscopy and stoppedflow optical spectroscopy. For the I100W mutant, a transient species is observed with an absorption maximum at 500 nm. EPR and Mossbauer spectra of RFQ samples identified this species as a diiron(III,IV) cluster spin-coupled to a neutral W radical. ENDOR spectra of this intermediate confirmed the protonation state and type of the amino acid radical and also identified a labile terminal water or hydroxide on the diiron center. / (cont.) Decay of this intermediate results in hydroxylation of the W radical. A diamagnetic precursor to the mixed-valent diiron(III,IV) center was also observed at an earlier time-point, with Mossbauer parameters typical of high-spin FeIII. We have tentatively assigned this antiferromagnetically-coupled diiron(III) intermediate as a peroxo-bridged cluster. A similar diiron(III) species is observed in RFQ Mossbauer samples from the reaction of reduced wild type hydroxylase with dioxygen. Substrate accelerates the decay rate of this species, providing evidence for the diiron(III) transient as the active oxidant. Under steady state conditions, hydrogen peroxide was generated in the absence of substrate. The oxidized hydroxylase also decomposed hydrogen peroxide to liberate dioxygen if no reducing equivalents were present. This catalase activity suggests that dioxygen activation could be reversible. The linear free energy relationship determined from steady state hydroxylation of para substituted phenols has a negative slope. A value of ? < 0 is indicative of electrophilic attack on the aromatic substrate by the oxidizing diiron(III) intermediate. The results from these steady state and pre-steady experiments provide compelling evidence that the diiron(III) transient is the active oxidant in ToMO and is a peroxodiiron(III) transient, despite differences between the optical and Mossbauer spectroscopic parameters and those of other peroxodiiron(III) centers. Enzymatic oxidation of the radical clock substrate probe, norcarane, by ToMO gives rise to both desaturation and hydroxylation products, norcarenes and norcaranols respectively. / (cont.) Norcarenes are better substrates for this enzyme system than norcarane, producing additional oxidation products. In all, more than twenty oxidation products were characterized in these reaction mixtures, half of which arose from norcarene oxidation. Accounting for these secondary oxidation products, we determined that no substrate radical intermediates with a significant lifetime (t < 25 ps) are formed during catalysis. / by Leslie Justin Murray. / Ph.D.

Thesis (M.S.)--Massachusetts Institute of Technology, Dept. of Chemistry, 1997. / Includes bibliographical references (leaves 69-70). / by Sean Konrad McHugh. / M.S.