Abstract

The principal goal of this research is a fundamental molecular understanding of the active site structure and catalytic mechanism(s) of copper containing amine oxidases. These enzymes are involved in the metabolism of biogenic primary amines, which have a variety of physiological functions in the cardiovascular, gastrointestinal, and nervous systems; amine oxidases are also responsible for the crosslinking of connective tissue (elastin and collagen) in aorta, lung, and cartilage. It is proposed to exploit the spectroscopic properties of the Cu(II) ion as a built in probe of the active site in amine oxidases purified from several sources. Variable temperature absorption, circular dichroism, magnetic circular dichroism, and emission spectroscopy as well as resonance raman and X-ray absorption spectroscopy will be employed. These spectroscopic techniques will also be used to determine how the Cu(II) interacts with any additional cofactor present in amine oxidases. Data will be obtained on the native, reduced, and copper-depleted forms of the enzymes. Effects of inhibitors, substrates, and substrate analogs on the spectra will be measured. Metal-substituted enzymes will be prepared to afford additional spectroscopic probes and to directly test possible metal ion roles in catalysis.

  Funding Agency

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM027659-06
Application #
3274863
Study Section
Physical Biochemistry Study Section (PB)
Project Start
1980-07-01
Project End
1988-06-30
Budget Start
1985-07-01
Budget End
1986-06-30
Support Year
6
Fiscal Year
1985
Total Cost
Indirect Cost

  Institution

Name
Amherst College
Department
Type
Schools of Arts and Sciences
DUNS #
City
Amherst
State
MA
Country
United States
Zip Code

  Publications

Cowley, Ryan E; Cirera, Jordi; Qayyum, Munzarin F et al. (2016) Structure of the Reduced Copper Active Site in Preprocessed Galactose Oxidase: Ligand Tuning for One-Electron O2 Activation in Cofactor Biogenesis. J Am Chem Soc 138:13219-13229
Liu, Yi; Mukherjee, Arnab; Nahumi, Nadav et al. (2013) Experimental and computational evidence of metal-O2 activation and rate-limiting proton-coupled electron transfer in a copper amine oxidase. J Phys Chem B 117:218-29
Mills, Stephen A; Brown, Doreen E; Dang, Kaitlyn et al. (2012) Cobalt substitution supports an inner-sphere electron transfer mechanism for oxygen reduction in pea seedling amine oxidase. J Biol Inorg Chem 17:507-15
Rokhsana, Dalia; Howells, Alta E; Dooley, David M et al. (2012) Role of the Tyr-Cys cross-link to the active site properties of galactose oxidase. Inorg Chem 51:3513-24
McGrath, Aaron P; Hilmer, Kimberly M; Collyer, Charles A et al. (2010) A new crystal form of human diamine oxidase. Acta Crystallogr Sect F Struct Biol Cryst Commun 66:137-42
Ran, Yanchao; Liu, Mengyao; Zhu, Hui et al. (2010) Spectroscopic identification of heme axial ligands in HtsA that are involved in heme acquisition by Streptococcus pyogenes. Biochemistry 49:2834-42
McGrath, Aaron P; Hilmer, Kimberly M; Collyer, Charles A et al. (2009) Structure and inhibition of human diamine oxidase. Biochemistry 48:9810-22
Rokhsana, Dalia; Dooley, David M; Szilagyi, Robert K (2008) Systematic development of computational models for the catalytic site in galactose oxidase: impact of outer-sphere residues on the geometric and electronic structures. J Biol Inorg Chem 13:371-83
Langley, David B; Brown, Doreen E; Cheruzel, Lionel E et al. (2008) Enantiomer-specific binding of ruthenium(II) molecular wires by the amine oxidase of Arthrobacter globiformis. J Am Chem Soc 130:8069-78
Zhu, Hui; Xie, Gang; Liu, Mengyao et al. (2008) Pathway for heme uptake from human methemoglobin by the iron-regulated surface determinants system of Staphylococcus aureus. J Biol Chem 283:18450-60

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