UM Biophysics Faculty | David P Ballou

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David P Ballou

Professor - Biological Chemistry

David Ballou

M.S., Ph.D., University of Michigan

Dept: Department of Biological Chemistry

Office Address: 5420 Med Sci I

Phone: (734) 764-9582

Email: dballou@umich.edu

Ballou group home page

Our laboratory is studying a) how O₂ is activated by various oxygenases, b) mechanisms of electron transfer in proteins, and c) how redox reactions are controlled in biology. Redox enzymes participate in all aerobic organisms and are crucial to the normal metabolism of of endogenous substrates, as well as to the metabolism of pharmacological and toxic substances. We are examining flavoproteins, hemoproteins, and nonheme iron proteins, expecially those participating as oxygenases. In addition, other redox proteins that can be isolated in homogeneous form are being investigated. Our goal is to learn in detail how a few of these enzymes, which serve as prototypes for the flavin, heme, and nonheme iron classes of enzymes, activate oxygen and control reactions with organic compounds.

Many flavin-containing hydroxylases participate in the degradation of aromatic compounds in the soil. We use physical techniques such as rapid reaction spectroscopy to determine properties of these hydroxylases. The figures below illustrate how spectra of 4a-flavin hydroperoxide and 4a-flavin hydroxide intermediates can be captured by stopped-flow rapid mixing techniques.

Nonheme iron-containing dioxygenases and monooxygenases found in soil bacteria are the starting points for activating and utilizing persistent and toxic aromatic and aliphatic compounds in the environment. We are investigating phthalate dioxygenase, which is a prototype for one of the classes of nonheme iron oxygenases. The stability and availability of this enzyme system has allowed us to carry out detailed spectroscopic, crystallographic, and biochemical studies of this system. This system includes several redox-reactive sites including FMN, a plant-type [2Fe-2S] center, a Rieske [2Fe-2S] center, and a mononuclear Fe²⁺ center where oxygenation takes place.

Cytochrome P450 enzymes are involved ubiquitously in biochemical reactions including drug metabolism, regulation, hormone synthesis, and biodegradation. We have been characterizing intermediates thought to be involved in the reactions of P450. We have developed conditions for observing Cpd I, Cpd II, and Cpd ES, and are characterizing their properties by UV-vis, EPR, and Mossbauer spectroscopy. We are also examining how the protein environment controls flavins to elicit a wide variety of chemical reactions. This includes regulating the redox potential, providing (or preventing access to) protons, and physically creating the proper binding, room to move, or substrate access.

Data Plot

Representative Publications

Ortiz-Maldonado, M., Ballou, D.P., and Massey, V.: Use of Free Energy Relationships to Probe the Individual Steps of Hydroxylation of p-Hydroxybenzoate Hydroxylase: Studies with a Series of 8-Substituted Flavins, Biochemistry, 1999, 38, 8124-8137.

Coulter, E.D., Moon, N., Batie, C.J., Dunham, W.R., and Ballou, D.P.: Electron Paramagnetic Resonance Measurements of the Ferrous Mononuclear Site of Phthalate Dioxygenase Substituted with Alternate Metal Ions: Direct Evidence for Ligation of Two Histidines in the Copper(II)-Reconstituted Protein, Biochemistry, 1999, 38, 11062-11072.

Wang, J., Ortiz-Maldonado, M., Entsch, b., Massey, V., Ballou, D., Gatti, D.L.: Protein and Ligand Dynamics in 4-Hydroxybenzoate Hydroxylase, Proc. Natl. Acad. Sci., 2002, 99, 608-613.

Ortiz-Maldonado M, Entsch B, Ballou DP.: Conformational changes combined with charge-transfer interactions are essential for reduction in catalysis by p-hydroxybenzoate hydroxylase. Biochemistry 2003 Sep 30;42(38):11234-42.

Bauer H, Massey V, Arscott LD, Schirmer RH, Ballou DP, Williams CH Jr.: The mechanism of high Mr thioredoxin reductase from Drosophila melanogaster. J Biol Chem. 2003 278, 33020-8.

Gromer S, Johansson L, Bauer H, Arscott LD, Rauch S, Ballou DP, Williams CH Jr, Schirmer RH, Arner ES.: Active sites of thioredoxin reductases: Why selenoproteins? Proc Natl Acad Sci USA 2003, 100, 12618-12623.

Ninfa, A. and Ballou, D.P.: Fundamental Laboratory Approaches for Biochemistry and Biotechnology (1998) Fitzgerald Science Press, Inc., Bethesda, 1998.

Regeimbal J, Gleiter S, Trumpower BL, Yu CA, Diwakar M, Ballou DP, Bardwell JC. Disulfide bond formation involves a quinhydrone-type charge-transfer complex. Proc Natl Acad Sci U S A. 2003, 100, 13779-13784.

Xia L, Ballou DP, Marsh EN. Role of arg100 in the active site of adenosylcobalamin-dependent glutamate mutase. Biochemistry. 2004, 43, 3238-45.

Entsch, B, Cole, LJ, and Ballou, DP, Protein dynamics and electrostatics in the function of p-hydroxybenzoate hydroxylase. Arch Biochem Biophys. 2005 43: 297-311.

Tarasev, M and Ballou, DP, Rates of the phthalate dioxygenase reaction with oxygen are dramatically increased by interactions with phthalate and phthalate oxygenase reductase.
Biochemistry. 2004 43 :12799-808.

Glascock, M.C., Ballou, D.P., and Dawson, J.H.: Effector Role of Reduced Putidaredoxin on the Oxygenation Reaction of Cytochrome P450-CAM; Perturbed Oxyferrous Form, a New Intermediate in the Reaction. J. Biological Chemistry, 2005, 280, 42134-42141.

Grzyska PK, Ryle MJ, Monterosso GR, Liu J, Ballou DP, Hausinger R.P. Steady-State and Transient Kinetic Analyses of Taurine/alpha-Ketoglutarate Dioxygenase: Effects of Oxygen Concentration, Alternative Sulfonates, and Active-Site Variants on the Fe(IV)-oxo Intermediate. Biochemistry 2005, 44, 3845-3855.

Spolitak T, Dawson JH, Ballou DP. Reaction of ferric cytochrome P450CAM with peracids: Kinetic characterization of intermediates on the reaction pathway. J Biol Chem. 2005, 280, 20300-20309.

Ballou, D.P., Entsch, B., Cole, L.J., Dynamics involved in catalysis by single-component and two-component flavin-dependent aromatic hydroxylases. Biochem. Biophys. Res. Commun. 2005, 338, 590-598.

Brender, J.R., Dertouzos, J., Ballou, D.P., Massey, V., Palfey, B.A., Entsch, B., Steel, D.G., Gafni, A. Conformational Dynamics of the Isoalloxazine in Substrate-free p-Hydroxybenzoate Hydroxylase: Single-Molecule Studies. J. American Chemical Society, 2005, 127, 18171-18178.

PubMed Listing of Ballou Publications

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Biophysics Research Division University of Michigan 4028 Chemistry Building 930 North University Avenue Ann Arbor, MI 48109-1055 UMsitemaker	Phone: (734) 763-6722 Fax: (734) 764-3323 E-mail: biophysics@umich.edu Last Updated: 1/26/2006 © Copyright 2006 University of Michigan