This project address the structure and function of enzymes involved in the degradation of polycyclic aromatic and haloaromatic compounds. A number of these compounds, such as the PCBs and dioxins, are well-known as persistent pollutants of health and environmental significance. X-ray crystallography will be used to determine high resolution structures of various states of enzymes in the PCB degradative pathway. The extradiol dioxygenase 2,3-dihydroxybiphenyl 1,2-dioxygenase catalyzes the key step of ring cleavage. To test mechanistic proposals, structural studies of the enzyme from Pseudomonas LB400 will be extended to several binary and ternary complexes analogous to proposed intermediates. Site-directed variants of residues believed to be of catalytic significance will also be examined. Additional studies will evaluate the structural origins of catalytic impasses related to substrate preference and metabolite inhibition. The crystal structure of a second extradiol dioxygenase that expresses important differences in substrate preference as well as resistance to metabolite inhibition will be determined and analyzed. This second enzyme belongs to a different structural class and is of general interest with respect to the structural evolution of proteins. Structural studies will be initiated for components of a second key enzyme from this pathway, the ring-hydroxylating dioxygenase biphenyl 2,3-dioxygenase. In this case, one goal is to analyze the structure of the Rieske component with respect to structural features that influence its distinct redox properties. The proposed studies are part of an international effort that joins our structural investigations with the complementary research of others. The general goals of this effort are: to understand the mechanisms of the enzymes involved in the biphenyl-PCB pathways; to understand and alter substrate preferences at key steps in the pathways; to understand and remove catalytic impasses such as the irreversible inhibition of dioxygenases by chlorinated metabolites; and, in a general sense, to exploit the results of basic research along a variety of fronts in the development of microbial systems with an enhanced ability to degrade a broad range of aromatic and haloaromatic pollutants.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM052381-07
Application #
6386131
Study Section
Biophysical Chemistry Study Section (BBCB)
Program Officer
Lewis, Catherine D
Project Start
1995-05-08
Project End
2004-04-30
Budget Start
2001-05-01
Budget End
2004-04-30
Support Year
7
Fiscal Year
2001
Total Cost
$194,468
Indirect Cost
Name
Purdue University
Department
Biology
Type
Schools of Arts and Sciences
DUNS #
072051394
City
West Lafayette
State
IN
Country
United States
Zip Code
47907