- 3097B Shelby Hall
- (205) 348-0269
- (205) 348-9104
Metal-related diseases, Protein structure-function studies, Amide hydrogen-deuterium exchange mass spectrometry. Research in the Busenlehner group is centered on the study of proteins that are involved in metal-related diseases and conditions. This research utilizes many classical biochemical, biophysical and bioinorganic methods to study the binding of native and non-native metal ions to physiologically relevant proteins. Amide proton hydrogen-deuterium exchange mass spectrometry is a structural tool that is used quite extensively in the Busenlehner group. Amide proton exchange is a sensitive technique primarily used to probe the functional importance of structural elements and dynamic motions in proteins. We are using the technique to localize metal-binding and protein-protein interaction sites, as well as to map any conformational changes that result.
- Structural and Functional Investigations of the Iron Chaperone Frataxin. Many human diseases are the result of imbalances in native metal ion homeostatic mechanisms. In the case of the neurodegenerative disease Friedreich’s ataxia (FRDA), insufficient levels of the iron chaperone frataxin result in mitochondrial iron accumulation, but not in a bioavailable form. FRDA is a genetic disease marked by cardiomyopathy, cardiac iron deposits, degeneration of sensory neurons and early death. In the mitochondria, iron uptake and transport must be tightly controlled. Unbound iron can interact with free radicals generated from energy production, which leads to oxidative stress. By studying the iron chaperone function of frataxin, a piece of the puzzle as to how iron is trafficked in the mitochondria and is inserted into specific metalloproteins and enzymes can be filled in. The project seeks to answer structural and mechanistic questions about the biology and bioinorganic chemistry of frataxin as it relates to FRDA.
- Characterizing Metalloestrogen Interactions with Estrogen Receptor-a. Exposure to lead, cadmium and arsenic is associated with increased risk for a variety of cancers. The underlying mechanisms of carcinogenesis, and even toxicity, are still unclear. Metal ions can act as estrogen analogs (metalloestrogens) and interaction and activate with estrogen receptor-a. Amide H/D exchange will be used to localize ERa binding site(s) for metalloestrogens. These and other experiments will address how the binding of specific metal ions can conformationally mimic estradiol-based activation of ERa and will reveal if reasonable physiological concentrations of specific metal ions are capable of non-steroidal estrogen receptor activation.
- Heavy Metal Inactivation of DNA Repair Proteins. There is increasing evidence that metallo-inhibition of DNA repair proteins is involved in metal-induced carcinogenesis. Many, but not all of these proteins rely on native metal ions for their structure or function. This project will study the equilibrium binding of heavy metals to E. coli and human DNA repair proteins and correlate metal binding constants and coordination environment with the ability to recognize synthesized DNA templates with appropriate lesions. Metal binding inactivation will also be studied structurally using H-D exchange mass spectrometry.
“Structural elements involved in proton translocation by cytochrome c oxidase as revealed by backbone amide hydrogen-deuterium exchange of the E286H mutant.” Busenlehner, L. S.; Braenden, G.; Namslauer, I.; Brzezinski, P.; Armstrong, R. N. Biochemistry 47, 73-83 (2008).
“Location of Substrate Binding Sites within the Integral Membrane Protein Microsomal Glutathione Transferase-1.” Busenlehner, L.S., ¿lander, J., Jegerscˆhld, C., Bhakat, P., Hebert, H., Morgenstern, R., and Armstrong, R.N. Biochemistry 46: 2812-2822 (2007).
“Mapping Protein Dynamics in Catalytic Intermediates of the Redox-Driven Proton Pump Cytochrome c Oxidase.” Busenlehner, L.S., Salomonsson, L., Brzezinski, P., and Armstrong, R.N. Proc. Nat. Acad. Sci. 103, 15398-15403 (2006).
“Kinetics of Metal Binding by the Toxic Metal-Sensing Transcriptional Repressor Staphylococcus aureus pI258 CadC.” Busenlehner, L.S. and Giedroc, D.P. J. Inorg. Biochem. 100, 1024-1034 (2006).
“Insights into Enzyme Structure and Dynamics Elucidated by Amide H/D Exchange Mass Spectrometry.” Busenlehner, L.S. and Armstrong, R.N. Arch. Biochem. Biophys. 433, 34-46 (2005).
“Stress Sensor Triggers Conformational Response of the Integral Membrane Protein Microsomal Glutathione Transferase 1.” Busenlehner, L.S., Codreanu, S.G., Holm, P.J., Bhakat, P., Hebert, H., Morgenstern, R., and Armstrong, R.N. Biochemistry 43, 11145-11152 (2004).
“Ratiometric Pulsed Alkylation-Mass Spectrometry as a Probe of Thiolate Reactivity in Different Metalloderivatives of S. aureus pI258 CadC.” Apuy, J.L., Busenlehner, L.S., Russell, D.H., and Giedroc, D.P. Biochemistry 43, 3824-3834 (2004).
“The SmtB/ArsR Family of Metalloregulatory Transcriptional Repressors: Structural Insights into Prokaryotic Metal Resistance.” Busenlehner, L.S., Pennella, M.A., and Giedroc, D.P. FEMS Microbiol. Rev. 27, 131-143 (2003).
“Elucidation of Primary (a3N) and Vestigial (a5) Heavy Metal Binding Sites in S. aureus pI258 CadC: Evolutionary Implications for Metal Ion Selectivity of ArsR/SmtB Metal Sensor Proteins.” Busenlehner, L.S., Wang, T.C., Penner-Hahn, J.E., and Giedroc, D.P. J. Mol. Biol. 319, 685-701 (2002).
“Characterization of a Metalloregulatory Bismuth(III) Site in Staphylococcus aureus pI258 CadC Repressor.” Busenlehner, L.S., Apuy, J.L., and Giedroc, D.P. J. Biol. Inorg. Chem. 7, 551-559 (2002).