Browsing by Subject "Decarboxylation"
Now showing 1 - 3 of 3
- Results Per Page
- Sort Options
Item Mutagenic analysis of the decarboxylases and hydratases in parallel meta-fission pathways(2008-08) Miller, Scott Garrett; Whitman, Christian P.The catechol meta-fission pathway, a degradation pathway for simple aromatic compounds, is rich in enzyme chemistry and replete with structural and evolutionary diversity. Vinyl pyruvate hydratase (VPH) and MhpD catalyze the same reaction in this pathway, but in different bacterial species. These metal ion-dependent enzymes reportedly catalyze a 1,5-keto-enol tautomerization reaction followed by a Michael addition of water. MhpD, and most likely VPH, are members of the fumarylacetoacetate hydrolase (FAH) superfamily. The crystal structure of MhpD and the sequence of VPH identified four potential active site residues, Lys-60, Leu-72, Asp-78, and Ser-160 (Ser-161 in VPH). The K60A and D78N mutants of VPH and MhpD had the most damaging effects on catalysis. Moreover, the K60A mutant seemingly uncoupled tautomerization from hydration and provided evidence for an [alpha, beta]-unsaturated ketone in the reaction. The effects of the L72A and S160A (S161A in VPH) mutants were smaller, suggesting less important roles in the mechanism. 5-(carboxymethyl)-2-Oxo-3-hexene-1,6-dioate decarboxylase (COHED) is a metal ion-dependent enzyme in the homoprotocatechuate (HPC) pathway, a chromosomally encoded meta-fission pathway from Escherichia coli C that parallels the catechol meta-fission pathway. COHED is also a member of the FAH superfamily. It is a monomeric protein with two domains. It is postulated that the C-terminal domain catalyzes the decarboxylation reaction and the N-terminal domain carries out the 1,3-keto-enol tautomerization reaction. Site-directed mutagenesis, NMR, and kinetic analysis with different substrates and inhibitors have identified three potential active-site residues Glu-276, Glu-278 (in the C-terminal domain), and Lys-110 (in the N-terminal domain). Replacement of either glutamate with a glutamine eliminated both the decarboxylase and tautomerase activities. The K110A mutant also diminished both activities, but more importantly eliminated the C-3 proton/deuteron exchange reaction observed for substrate analogs. The enzymes of the catechol and homoprotocatechuate pathways provide examples of enzyme optimization toward a specific substrate even among related compounds, as reflected by the FAH superfamily. Hence, the results of these studies add to the growing body of information about how enzymes evolve and how pathways are assembled.Item Structural study of a kero base of formula C₁₆H₂₅N(1934) Lackey, Robert Woodfin, 1899-; Bailey, James RobinsonItem Synthesis of top coat surface treatments for the orientation of thin film block copolymers(2013-08) Chen, Christopher Hancheng; Willson, C. Grant, 1939-Block copolymer self-assembly has demonstrated sub-optical lithographic resolution . High values of chi, the block copolymer interaction parameter, are required to achieve next-generation lithographic resolution . Unfortunately, high values of chi can lead to thin film orientation control difficulties , which are believed to be caused by large differences in the surface energy of each block relative to the substrate and the free surface. The substrate-block interface can be modified to achieve a surface energy intermediate to that of each individual block ; the air-polymer interface, however, presents additional complications. This thesis describes the synthesis of polymers for top coat surface treatments, which are designed to modify the surface energy of the air-block copolymer interface and enable block copolymer orientation control upon thermal annealing. Polymers with β-keto acid functionality were synthesized to allow polarity switching upon decarboxylation. Syntheses of anhydride containing polymers were established that provide another class of polarity switching materials.