WALKER, ANN F. / Quimica / Investigadores

Centro Internacional de Investigación Científica


Quimica / WALKER, ANN F.



Regents Professor Emerita - Retired
Department of Chemistry, The University of Arizona, Tucson, Arizona, US.

Porphyrins, metalloporphyrins, metallocorrins, chlorins, glyoximes, phthalocyanines and other related macrocycles. Gene synthesis and bacterial expression of heme proteins; 1- and 2-dimensional NMR spectroscopy, structures and electron transfer reactions of metalloproteins; cytochromes; nitrosylheme proteins; binding of small molecules to heme proteins; metal nucleus NMR spectroscopy, continuous wave and pulsed EPR spectroscopy, electronic, low temperature MCD, Mössbauer and PES spectroscopies, and electrochemistry of transition metal complexes; thermodynamics and kinetics of ligand addition to metal complexes; electronic properties of ligands; porphyrins and metalloporphyrins as medical imaging agents.

Her research group studies a wide range of bioinorganic and biological systems, all of which fall under the general theme of gaining a better understanding of the heme centers in heme proteins that are vital to the life of almost all living organisms.

The overall goals of this research are :

1. To characterize the nitrosylheme proteins from blood-sucking insects. Nitric oxide has been shown to be an important neurotransmitter, vasodilator, and other chemical messenger. We have recently shown that Rhodnius prolixus (the “kissing bug”)1,2 and Cimex lectularius (the bedbug)3 each have at least one NO-carrying heme protein in their saliva that helps them succeed in their goal of living on the blood of higher animals. In collaboration with the research group of Dr. William Montfort, Dept. of Biochemistry, and others off-campus, we are investigating the 3-D structures, spectroscopy (NMR, EPR, UV-vis, MCD, Mössbauer and resonance Raman), thermodynamics (kinetics and equilibria of NO binding, and reduction potentials in the absence and presence of NO) of the NO-binding heme proteins from both of these insects. Current research on these proteins includes preparation and investigation of appropriate site-directed mutants to test hypotheses as to which amino acid side chains affect NO and histamine binding and release.

2. To prepare and investigate site-directed mutants of the membrane-bound cytochrome b of the bc1 complex (respiratory Complex III) of the photosynthetic bacterium Rhodobacter sphaeroides. The mutants are chosen so as to interfere with the binding of one of the histidine ligands of either the high- or the low-potential heme of cytochrome b, in order to determine how this interference affects the EPR spectrum and the reduction potential of that heme, and the rate of electron transfer through the low-potential half of the bifurcated electron transfer pathway of the Q-cycle.

3. To investigate the reaction mechanisms, substrate specificity and domain interactions of cytochrome P450BM3 and several insect cytochromes P450. Cytochromes P450 are monooxygenase enzymes that comprise about 10% of the protein found in the human liver. They hydroxylate or epoxidize a wide range of substrates including (but not limited to) steroid hormones and xenobiotics. P450BM3, from Bacillus megaterium, has been described as a “complete enzyme” because in one polypeptide it has both the heme and reductase domains necessary to catalyze the oxidation of substrates in the presence of molecular oxygen and NADPH, and its reactions are 100-1000 times faster than those of reconstituted microsomal P450s. We are using the “scanning chimeragenesis” approach to create P450BM3 mutants with altered substrate specificity. Techniques being used include genetic engineering, protein expression and purification, optical, multidimensional NMR, EPR and pulsed EPR spectroscopies, stopped-flow kinetics, gas chromatography and mass spectrometry (for identification of the products of enzymatic reactions).

4. To investigate model hemes and heme proteins by multidimensional NMR spectroscopy. The goal of these studies is to use (and to further develop) modern NMR techniques, such as COSY, NOESY, TOCSY, ROESY, HOESY, HMQC, HMBC, etc., to determine the solution structure and investigate the dynamic reactions (including axial ligand rotation, macrocycle inversion and ligand exchange) of selected model heme complexes in which the oxidation state of iron ranges from +1 to +4, all of which have relevance to the biological roles of hemes and heme proteins. Computer modeling of the structural and dynamic features of these molecules will continue to complement the studies.

- DuPont Teaching Fellowship, Brown University, 1965-66
- NIH Postdoctoral Fellowship, University of California, Los Angeles, 1966-67
- NATO Senior Research Fellow in Science, University of Kent, Canterbury, U.K., Fall, 1974
- NIH Research Career Development Award, 1976-81
- Fulbright Visiting Scholar (Travel Grant), Technical University of Munich, Physics Department, Spring, 1977
- Visiting Professor, University of Queensland, Australia, Spring, 1984
- NIH National Research Service Award, Spring, 1984
- Fellow, American Association for the Advancement of Science, 1984
- Meritorious Performance and Professional Promise Award, SFSU*, 1985, 1986, 1988
- California State University Research Leave Award, SFSU*, Spring, 1989
- Mortar Board Certificate of Recognition, University of Arizona, 1996
- Invité, Université de Rennes, October, 1996
- Associate Editor, J. Am. Chem. Soc., March, 1998-Present
- Recipient, Francis P. Garvan-John M. Olin Medal, American Chemical Society, March, 2000. View Photos of the award ceremonies.
- Inducted as Regents Professor, Department of Chemistry, University of Arizona, February, 2001.
- Recipient, Luigi Sacconi Medal, Inorganic Chemistry Division, Italian Chemical Society, 2001; awarded 2002
- Alexander von Humboldt Senior Research Award in Science, 2003-04
- Alfred Bader Award in Bioinorganic Chemistry, American Chemical Society, March, 2006.

\"Effect of the N-Terminus on Heme Cavity Structure, Ligand Equilibrium and Rate Constants, and Reduction Potentials of Nitrophorin 2 from Rhodnius prolixus,\" Berry, R. E.; Shokhireva, T. Kh.; Filippov, I.; Shokhirev, M. N.; Zhang, H.; Walker, F. A. Biochemistry 2007, 46, 6830-6843. (DOI: 10.1021/bi7002263)

Effect of Mutation of Carboxylate Side-Chain Amino Acids Near the Heme on the Midpoint Potentials and Ligand Binding Constants of Nitrophorin 2 and its NO, Histamine and Imidazole Complexes,\" Berry, R. E.; Shokhirev, M. N.; Ho, A. Y. W.; Yang, F.; Shokhireva, T. K.; Zhang, H.; Weichsel, A.; Montfort, W. R.; Walker, F. A. J. Am. Chem. Soc. 2009, 131, 2313-2327. (DOI: 10.1021/ja00006a066)

\"1H and 13C NMR Spectroscopic Studies of the Ferriheme Resonances of the Low- Spin Imidazole, Histamine and Cyanide Complexes of wt Nitrophorin 2 and NP2(V24E) as a Function of pH,\" Yang, F.; Knipp, M.; Berry, R. E.; Shokhireva, T. K.; Zhang, H.; Walker, F. A. J. Biol. Inorg. Chem. 2009, 14, 1077-1095. (DOI: 10.1007/200775-009-00551-3)

“The Unprecedented Peroxidase-like Activity of Rhodnius prolixus Nitrophorin 2: Identification of the [FeIV=O Por·]+ and [FeIV=O Por](Tyr38·) Intermediates and Their Role(s) in Substrate Oxidation,” Singh, R.; Berry, R. E.; Yang, F.; Zhang, H.; Walker, F. A.; Ivancich, A. I. Biochemistry 2010, 49, 8857-8872.
\"NMR Studies of Nitrophorin Distal Pocket Side Chain Effects on the Heme Orientation and Seating of NP2 as Compared to NP1,\" Shokhireva, T. K.; Berry, R. E.; Zhang, H.; Walker, F. A. J. Inorg. Biochem. 2011, 105, 1238-1257. (DOI: 10.1016/j.jinorgbio.2011.06.001) PMCID: PMC3155619

\"Nuclear Inelastic Scattering and Mössbauer Spectroscopy as Local Probes for Ligand Binding Modes and Electronic Properties in Proteins: Vibrational Behavior of a Ferriheme Center Inside a ?-Barrel Protein,\" Moeser, B.; Janoschka, A.; Wolny, J. A.; Paulsen, H.; Filippov, I.; Berry, R. E.; Zhang, H.; Chumakov, A. I.; Walker, F. A.; Schünemann, V. J. Am. Chem. Soc. 2012, 134, 4216-4228. (DOI: 10.1021/ja210067t) PMCID: PMC3296872

\"Native N-Terminus Nitrophorin 2 from the Kissing Bug: Similarities to and Differences from NP2(D1A),\" Berry, R. E.; Muthu, D.; Garrett, S. A.; Shokhireva, T. K.; Zhang, H.; Walker, F. A. Chem. Biodiv. 2012, 9, 1739-1755. (DOI: 10.1002/cbdv.201100449) PMCID:PMC3523740

\"Electron Spin Density on the Axial His Ligand of High-Spin and Low-Spin Nitrophorin 2 Probed by Heteronuclear NMR Spectroscopy,\" Abriata, L. A.; Zaballa M-E.; Berry, R. E.; Yang, F.; Zhang, H.; Walker, F. A.; Vila, A. J. Inorg. Chem. 2013, 52, 1285-1295. (DOI: 10.1021/ic301805y)

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