The long term research goals of this program are to attain a molecular level understanding of the remarkable functional diversity of heme proteins, substances that are of critical importance in biological systems, in general, and in human physiology in particular. In participating in and facilitating many critical biochemical processes, including oxygen and electron transport, as well as the elimination of internally generated and environmentally accumulated toxins, the inherent reactivity of the heme sites are effectively regulated by intermolecular interactions with the associated polypeptides and further controlled by intermolecular interactions with other proteins and small regulatory molecules. In addition to the more stable initial and final states, a great deal of interest is attached to the fleeting, but functionally important, transient or intermediate states.The essential strategy of this research program is to apply powerful spectroscopic probes, especially resonance Raman and time-resolved resonance Raman techniques, to the native and strategically manipulated proteins in order to reveal the molecular basis for this modulation of heme group reactivity. In fact, it is often the case that these particular methods are uniquely effective probes of these sites. The specific systems to be addressed in the present proposal are three important classes of heme proteins involved in mammalian physiology; cytochromes P450, which catalayze the conversion of xenobiotics, preparing them for elimination; guanylate cyclase, the most important receptor molecule for the neurotransmitter, NO, which serves a crucial regulatory function in human physiology; and, finally, a group of enzymes known as peroxidases that play a critical role in human defence against invading microorganisms.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK035153-19
Application #
7048471
Study Section
Metallobiochemistry Study Section (BMT)
Program Officer
Sechi, Salvatore
Project Start
1984-07-01
Project End
2008-03-31
Budget Start
2006-04-01
Budget End
2007-03-31
Support Year
19
Fiscal Year
2006
Total Cost
$238,813
Indirect Cost
Name
Marquette University
Department
Chemistry
Type
Schools of Arts and Sciences
DUNS #
046929621
City
Milwaukee
State
WI
Country
United States
Zip Code
53201
Mak, Piotr J; Denisov, Ilia G; Grinkova, Yelena V et al. (2011) Defining CYP3A4 structural responses to substrate binding. Raman spectroscopic studies of a nanodisc-incorporated mammalian cytochrome P450. J Am Chem Soc 133:1357-66
Mak, Piotr J; Zhang, Haoming; Hollenberg, Paul F et al. (2010) Defining the structural consequences of mechanism-based inactivation of mammalian cytochrome P450 2B4 using resonance Raman spectroscopy. J Am Chem Soc 132:1494-5
Balakrishnan, Gurusamy; Zhao, Xiaojie; Podstawska, Edyta et al. (2009) Subunit-selective interrogation of CO recombination in carbonmonoxy hemoglobin by isotope-edited time-resolved resonance Raman spectroscopy. Biochemistry 48:3120-6
Balakrishnan, Gurusamy; Ibrahim, Mohammed; Mak, Piotr J et al. (2009) Linking conformation change to hemoglobin activation via chain-selective time-resolved resonance Raman spectroscopy of protoheme/mesoheme hybrids. J Biol Inorg Chem 14:741-50
Rwere, Freeborn; Mak, Piotr J; Kincaid, James R (2008) Resonance Raman interrogation of the consequences of heme rotational disorder in myoglobin and its ligated derivatives. Biochemistry 47:12869-77
Denisov, Ilia G; Mak, Piotr J; Makris, Thomas M et al. (2008) Resonance Raman characterization of the peroxo and hydroperoxo intermediates in cytochrome P450. J Phys Chem A 112:13172-9
Mak, Piotr J; Kincaid, James R (2008) Resonance Raman spectroscopic studies of hydroperoxo derivatives of cobalt-substituted myoglobin. J Inorg Biochem 102:1952-7
Mak, Piotr J; Kaluka, Daniel; Manyumwa, Munyaradzi Edith et al. (2008) Defining resonance Raman spectral responses to substrate binding by cytochrome P450 from Pseudomonas putida. Biopolymers 89:1045-53
Mak, Piotr J; Im, Sang-Choul; Zhang, Haoming et al. (2008) Resonance Raman studies of cytochrome P450 2B4 in its interactions with substrates and redox partners. Biochemistry 47:3950-63
Mak, Piotr J; Denisov, Ilia G; Victoria, Doreen et al. (2007) Resonance Raman detection of the hydroperoxo intermediate in the cytochrome P450 enzymatic cycle. J Am Chem Soc 129:6382-3

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