Manganese redox chemistry is fundamental to biological processes at molecular, cellular and environmental scales, from antioxidant defense to oxygenic photosynthesis. Biomedically important enzymes including Mn superoxide dismutase (MnSOD), Mn catalase (MnC), and oxalate oxidase (OXO) require a catalytic manganese cofactor for their biological function, yet the molecular mechanisms involved in delivering the essential metal ion are just beginning to be studied in detail. The goal of this proposal is to advance our understanding of the biochemical mechanisms responsible for the metal-binding maturation of Mn metalloenzymes, including MnSOD. In vitro methods (including fluorescence monitored metal uptake kinetics) will be complemented by an analysis of processes and proteins required for metal binding by manganese metalloenzymes in vivo. This work is fundamental to understanding the role of manganese trafficking in human health and disease. Further, understanding the metal binding mechanism of the ubiquitous antioxidant metalloenzyme MnSOD may contribute new insight into biological defenses against oxidative stress, a key factor in aging and neurodegenerative disorders. This work may also shed light on diseases of metal homeostasis in which metal misincorporation occurs. Roles of manganese in pathogenesis will be explored through the expression and molecular characterization of manganese catalase homologs mined from the genome of a medically important microbial pathogen, the enterohaemorraghic E. coli O157:H7. Further, the role of high-valent manganese complexes in enzyme catalysis will be investigated by applying a combination of advanced spectroscopic methods and rapid kinetics to probe the active site chemistry of oxalate oxidase.

Public Health Relevance

Manganese is an element that plays many essential roles in health and disease. One of the important aims of this proposal is to investigate how living cells specifically deliver manganese to manganese-dependent enzymes so that they can perform their biological functions, providing important new insight into processes underlying aging and neurodegenerative disorders. The role of manganese in the virulence of the foodborne pathogen, E. coli O157:H7, will also be explored, and the mechanisms of manganese catalysis will be investigated in oxalate oxidase, an enzyme that is important in clinical bioanalytical chemistry.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
Research Project (R01)
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Macromolecular Structure and Function A Study Section (MSFA)
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Anderson, Vernon
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Oregon Health and Science University
Engineering (All Types)
Schools of Medicine
United States
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Whittaker, Mei M; Whittaker, James W (2014) Expression and purification of recombinant Saccharomyces cerevisiae mitochondrial carrier protein YGR257Cp (Mtm1p). Protein Expr Purif 93:77-86
Whittaker, James W (2013) Cell-free protein synthesis: the state of the art. Biotechnol Lett 35:143-52
McConnell, Iain L; Grigoryants, Vladimir M; Scholes, Charles P et al. (2012) EPR-ENDOR characterization of (17O, 1H, 2H) water in manganese catalase and its relevance to the oxygen-evolving complex of photosystem II. J Am Chem Soc 134:1504-12
Whittaker, Mei M; Lerch, Thomas F; Kirillova, Olga et al. (2011) Subunit dissociation and metal binding by Escherichia coli apo-manganese superoxide dismutase. Arch Biochem Biophys 505:213-25
Whittaker, James W (2010) Metal uptake by manganese superoxide dismutase. Biochim Biophys Acta 1804:298-307
Stich, Troy A; Whittaker, James W; Britt, R David (2010) Multifrequency EPR studies of manganese catalases provide a complete description of proteinaceous nitrogen coordination. J Phys Chem B 114:14178-88
Whittaker, Mei M; Whittaker, James W (2009) In vitro metal uptake by recombinant human manganese superoxide dismutase. Arch Biochem Biophys 491:69-74
Whittaker, Mei M; Whittaker, James W (2008) Conformationally gated metal uptake by apomanganese superoxide dismutase. Biochemistry 47:11625-36
Pan, Heng-Yen; Whittaker, Mei M; Bouveret, Romaric et al. (2007) Characterization of wheat germin (oxalate oxidase) expressed by Pichia pastoris. Biochem Biophys Res Commun 356:925-9
Whittaker, Mei M; Pan, Heng-Yen; Yukl, Erik T et al. (2007) Burst kinetics and redox transformations of the active site manganese ion in oxalate oxidase: implications for the catalytic mechanism. J Biol Chem 282:7011-23

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