The research outlined here is designed to achieve three main tasks: 1) Kinetic isotope effects (13C, 2H, and 3H) will be used to elucidate the structure of the intermediate that immediately follows the rate- determining step in the soybean lipoxygenase 1 (LOX-1) catalyzed oxidation of linoleic acid. 2) The effect of the identity of the transferred hydron (2H or 1H) upon secondary kinetic isotope effects (zH and 3H) will be studied using a method which allows for the simultaneous determination of all relevant isotope effects. 3) Finally, the effect steric perturbations near the active site upon the degree of tunneling of both 1H and 2H will be explored using a combination of site-directed mutagenesis, kinetic isotope effects, and a quantitative theoretical model. Completion of the first goal above promises to benefit research in drug design in the near term as human lipoxygenases have been implicated in asthma, athereosclerosis, and cancer. Attainment of the second goal will aid in overall understanding of the importance of secondary motions within the substrate during the primary tunneling event. Finally, completion of the third objective will serve to explain how conformational perturbations and internal motions near the active site affect tunneling. The last two objectives will confer long-term benefits upon the research community and could aid in the understanding of other enzymes which utilize tunneling.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Postdoctoral Individual National Research Service Award (F32)
Project #
1F32GM064218-01
Application #
6405572
Study Section
Physical Biochemistry Study Section (PB)
Program Officer
Ikeda, Richard A
Project Start
2001-09-03
Project End
Budget Start
2001-09-03
Budget End
2002-09-02
Support Year
1
Fiscal Year
2001
Total Cost
$33,260
Indirect Cost
Name
University of California Berkeley
Department
Chemistry
Type
Schools of Arts and Sciences
DUNS #
094878337
City
Berkeley
State
CA
Country
United States
Zip Code
94704
Meyer, Matthew P; Klinman, Judith P (2008) Synthesis of linoleic acids combinatorially-labeled at the vinylic positions as substrates for lipoxygenases. Tetrahedron Lett 49:3600-3603
Meyer, Matthew P; Tomchick, Diana R; Klinman, Judith P (2008) Enzyme structure and dynamics affect hydrogen tunneling: the impact of a remote side chain (I553) in soybean lipoxygenase-1. Proc Natl Acad Sci U S A 105:1146-51