Structural investigations of hydroxylase proteins and complexes in bacterial multicomponent monooxygenase systems

Bacterial multicomponent monooxgenases (BMMs) such as toluene/o-xylene monooxygenase (ToMO), phenol hydroxylase (PH), and soluble methane monooxygenase (sMMO) catalyze hydrocarbon oxidation reactions at a carboxylatebridged non-heme diiron center common to many systems in biology, as discussed in the first and subsequent chapters of this document. Chapter 1 provides a summary of various relationships between structure and activity in BMMs, as they have been determined through decades of research into BMM hydrocarbon catalysis. Presented in Chapter 2 are the structures of the native (ToMOH) and manganese(ll)-reconstituted (Mn(ll)-ToMOH) ToMO hydroxylase, at 1.85 A and 2.20 A resolution, respectively. The structure of Mn(ll)-ToMOH reveals an active site coordination and geometry similar to that in diferrous and manganese(ll)-reconstituted MMOH, indicating that it represents an analog of the diferrous ToMOH structure. Through comparison of the native ToMOH and Mn(II)-ToMOH structures, a collection of metal site oxidation state dependent conformational changes in conserved residues on the surface of the hydroxylase a-suibunit are observed, suggesting a relationship between active site oxidation state and component interactions in BMMs. Through analysis of the 1.85 A ToMOH structure, a series of hydrophobic cavities through the asubunit connecting the active site to the protein surface analogous to those previously noted in MMOH were also discovered as part of this work. Chapter 3 describes three X-ray crystal structures of ToMOH T201X mutants, and four structures of ToMOH N202X mutants at resolutions ranging from 1.90 to 2.90 A.

Department dc:contributor.department

Massachusetts Institute of Technology. Department of Chemistry

Grantor dc:publisher

Massachusetts Institute of Technology

Year dc:date.issued

2008

Author dc:creator

• McCormick, Michael S. (Michael Scott)

Advisor dc:contributor.advisor

• Stephen J. Lippard.

• Chemistry.

Statement dc:rights

• M.I.T. theses are protected by copyright. They may be viewed from this source for any purpose, but reproduction or distribution in any format is prohibited without written permission. See provided URL for inquiries about permission.

Licence dc:rights.uri

• http://dspace.mit.edu/handle/1721.1/7582

Language dc:language.iso

eng

Handle dc:identifier.uri

http://hdl.handle.net/1721.1/43769

OAI identifier oai:identifier

oai:dspace.mit.edu:1721.1/43769