Document Type

Dissertation

Degree

Doctor of Philosophy (PhD)

Department

Chemistry

First Advisor's Name

Xiaotang Wang

First Advisor's Committee Title

Committee Chair

Second Advisor's Name

Jose Almirall

Third Advisor's Name

Watson Lees

Fourth Advisor's Name

Wenzhi Li

Fifth Advisor's Name

Stanislaw Wnuk

Keywords

Chloroperoxidase, NMR, paramagnetic relaxation, molecular modeling, regioselective, enantioselective, biodegradation

Date of Defense

3-6-2013

Abstract

Chloroperoxidase (CPO) is the most versatile heme-containing enzyme that catalyzes a broad spectrum of reactions. The remarkable feature of this enzyme is the high regio- and enantio-selectivity exhibited in CPO-catalyzed oxidation reactions. The aim of this dissertation is to elucidate the structural basis for regio- and enantio-selective transformations and investigate the application of CPO in biodegradation of synthetic dyes.

To unravel the mechanism of CPO-catalyzed regioselective oxidation of indole, the dissertation explored the structure of CPO-indole complex using paramagnetic relaxation and molecular modeling. The distances between the protons of indole and the heme iron revealed that the pyrrole ring of indole is oriented toward the heme with its 2-H pointing directly at the heme iron. This provides the first experimental and theoretical explanation for the "unexpected" regioselectivity of CPO-catalyzed indole oxidation. Furthermore, the residues including Leu 70, Phe 103, Ile 179, Val 182, Glu 183, and Phe 186 were found essential to the substrate binding to CPO. These results will serve as a lighthouse in guiding the design of CPO mutants with tailor-made activities for biotechnological applications.

To understand the origin of the enantioselectivity of CPO-catalyzed oxidation reactions, the interactions of CPO with substrates such as 2-(methylthio)thiophene were investigated by nuclear magnetic resonance spectroscopy (NMR) and computational techniques. In particular, the enantioselectivity is partly explained by the binding orientation of substrates.

In third facet of this dissertation, a green and efficient system for degradation of synthetic dyes was developed. Several commercial dyes such as orange G were tested in the CPO-H2O2-Cl- system, where degradation of these dyes was found very efficient. The presence of halide ions and acidic pH were found necessary to the decomposition of dyes. Significantly, the results revealed that this degradation of azo dyes involves a ferric hypochlorite intermediate of CPO (Fe-OCl), compound X.

Identifier

FI13042318

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chapter 1

chap2.tex (59 kB)
chapter 2

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chapter 3

chap4.tex (35 kB)
chapter 4

chap5.tex (3 kB)
chapter 5

abstract.tex (2 kB)
abstract

prologue.tex (4 kB)
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reference

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vita.tex (1 kB)

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