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IN SILICO ENZYME MODELING:
A STUDY OF ENTROPIC CONTRIBUTIONS TO ENZYME CATALYSIS
AND THE DESIGN OF ARTIFICIAL KEMP ELIMINASE
by
Jie Cao
A Dissertation Presented to the
FACULTY OF THE USC GRADUATE SCHOOL
UNIVERSITY OF SOUTHERN CALIFORNIA
In Partial Fulfillment of the
Requirements for the Degree
DOCTOR OF PHILOSOPHY
(CHEMISTRY)
May 2011
Copyright 2011 Jie Cao
Object Description
| Title | In silico enzyme modeling: A study of entropic contributions to enzyme catalysis and the design of artificial kemp eliminase |
| Author | Cao, Jie |
| Author email | jiecao@usc.edu; caojiechem@gmail.com |
| Degree | Doctor of Philosophy |
| Document type | Dissertation |
| Degree program | Chemistry |
| School | College of Letters, Arts and Sciences |
| Date defended/completed | 2011-03-01 |
| Date submitted | 2011 |
| Restricted until | Restricted until 4 Nov. 2011. |
| Date published | 2011-11-04 |
| Advisor (committee chair) | Warshel, Ariehl |
| Advisor (committee member) |
Chen, Xiaojiang Gauderman, James |
| Abstract | Naturally occurring enzymes are exceptional catalysts. One of the most fundamental challenges in biotechnology, and in some respects in biochemistry, is the ability to design effective enzymes. Such ability would be one of the most convincing manifestations of a full understanding of the origin of enzyme catalysis. Thermodynamics properties play an important role in the catalytic effect. The free energy reflects entropic and enthalpic contributions, and understanding the origin of these contributions is undamentally important. It is important to evaluate the relative entropic and enthalpic contributions to the overall free energy. Such an evaluation is useful for assessing temperature effects and exploring specialized option in enzyme design.; Despite impressive progress, most of the advances on this front have been made by placing the reacting fragments in the proper places, rather than by optimizing the environment preorganization, which is the key factor in enzyme catalysis.; Improving preorganization would require approaches capable of evaluating reliably the actual catalytic effect. The empirical valence bond (EVB) method can give reliable prediction of enzyme activity and mutational effect.EVB is much less time-demanding and thus allows for the extensive conformational search needed in enzyme design. EVB can be used in the final screening stage in computer-aided enzyme design.; This work starts by looking thermodynamic properties of the enzymatic reactions for better understanding of general catalytic effects. It elucidates the microscopic nature of the contribution to the activation free energy and dissect its entropic and enthalpy components. This provides the physically based quantitative analysis of the temperature dependence of the entropy change in hydride transfer reactions. The analysis suggests that the observed effect is due to the response of the environment to the change in polarity of the reacting system upon moving from ground state to the transition state. Previously designed kemp eliminases as a benchmark for a computer aided enzyme design, using the EVB as the main screening tool. The observed absolute catalytic effect and the effect of directed; evolution are reproduced and analysed. It is found that, in the case of kemp eliminases, the transition state charge distribution makes it hard to exploit the active site polarity, even with the ability to quantify the effect of different mutations. Unexpectedly, it is found that the directed evolution mutants lead to the reduction of solvation of the reactant state by water molecules rather that to the more common mode of transition state stabilization used by naturally evolved enzymes.Finally it is pointed out that the difficulties in improving Kemp eliminase are due to the challenge in designing a preorganized environment that would exploit the small change it charge distribution during the formation of the transition state. |
| Keyword | computer-aided enzyme design; entropy; QM/MM; kemp eliminase; reorginization; EVB |
| Language | English |
| Part of collection | University of Southern California dissertations and theses |
| Publisher (of the original version) | University of Southern California |
| Place of publication (of the original version) | Los Angeles, California |
| Publisher (of the digital version) | University of Southern California. Libraries |
| Provenance | Electronically uploaded by the author |
| Type | texts |
| Legacy record ID | usctheses-m3892 |
| Rights | Cao, Jie |
| Repository name | Libraries, University of Southern California |
| Repository address | Los Angeles, California |
| Repository email | http://www.usc.edu/isd/libraries/services/ask_a_librarian/email/ |
| Filename | etd-cao-4380 |
| Archival file | uscthesesreloadpub_Volume29/etd-cao-4380.pdf |
Description
| Title | Page 1 |
| Full text | IN SILICO ENZYME MODELING: A STUDY OF ENTROPIC CONTRIBUTIONS TO ENZYME CATALYSIS AND THE DESIGN OF ARTIFICIAL KEMP ELIMINASE by Jie Cao A Dissertation Presented to the FACULTY OF THE USC GRADUATE SCHOOL UNIVERSITY OF SOUTHERN CALIFORNIA In Partial Fulfillment of the Requirements for the Degree DOCTOR OF PHILOSOPHY (CHEMISTRY) May 2011 Copyright 2011 Jie Cao |
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