cristina.timoteo

Cristina G. Timóteo
Post-doc Researcher
Collaborator
Research Group
Structural Molecular Biology
Chemistry Department, Lab. 425 / office 632A; Faculdade de Ciências e Tecnologia, UNL; Quinta da Torre; 2829-516 Caparica
- ext 10959
Research Interests
Biochemistry / EPR, UV-visible and Mössbauer Spectroscopies / Fast kinetics / Molecular Biology / Iron containing enzymes / Oxidative stress / Denitrification / Membrane-bound enzymes / Iron storage proteins / Oxidative stress sensors
Research work focused on detoxification mechanisms, connected to oxidative stress.
Study of enzymes catalysing elimination reactive oxygen species derived from molecular oxygen catalyst, isolated from denitrifying bacteria, including Pseudomonas stutzeri cytochrome c peroxidase and Pseudomonas nautica nitric oxide reductase. Determining conditions under which these enzymes are expressed by the bacteria, purification, equilibrium and fast kinetics, EPR, UV-visible, NMR and Mössbauer spectroscopies and molecular biology.
Establishment of the catalytic mechanism of iron storage and oxidation bacterial enzymes, such as Ferritins, Bacterioferritins or DNA protection enzymes from oxidative stress - DPS. Another enzyme containing an iron metal center, isolated from the plant Arabidopsis thaliana, and involved in resistance to climate change, was also the subject of study - a membrane-bound desaturase.
Study of an oxidative stress sensor - PerR – in a strict anaerobic bacterium - Desulfovibrio vulgaris. This bacterial repressor has a structural metal center with zinc and an active site containing iron. In the presence of these two metals and hydrogen peroxide it adopts an open conformation unable to bind to DNA, allowing the expression of oxidative stress protection enzymes, such as catalase or bacterioferritins.
More recently, study of a hemic protein, isolated from the bacteria Shewanella baltica, which binds molecular oxygen and has a highly unusual axial coordination.
Work has involved cloning and over-expressing the enzymes in Escherichia coli and application of spectroscopic techniques to caracterize the enzymes. To study the enzymatic reaction mechanism, UV-visible spectroscopy associated with stopped-flow technique has been used to identify coloured reaction intermediates as well as its formation and decay times. This information allows us to design Rapid-Freeze Quench experiments to produce samples for Mössbauer and EPR spectroscopies, which results in a more detailed analysis of intermediates and reaction mechanism.
Main publications
1. Cristina G. Timóteo, Márcia Guilherme, Daniela Penas, Filipe Folgosa, Pedro Tavares and Alice S. Pereira, “Desulfovibrio vulgaris bacterioferritin uses H2O2 as co-substrate for iron oxidation and reveals DPS-like DNA protection and binding activities”(2012) Biochem J, 446, 125-133.

2. Alice S. Pereira, Cristina G. Timóteo, Marcia Guilherme, Filipe Folgosa, Sunil G. Naik, Américo G. Duarte, Boi Hanh Huynh and Pedro Tavares, “Spectroscopic Evidence for and Characterization of a Trinuclear Ferroxidase Center in Bacterial Ferritin from Desulfovibrio vulgaris Hildenborough”, (2012) J Am Chem Soc, 134, 10822-10832.

3. Patrícia M. Paes de Sousa, David Rodrigues, Cristina G. Timóteo, Maria L. Simões Gonçalves, Graham W. Pettigrew, Isabel Moura, José J. G. Moura and Maria M. Correia dos Santos, “Analysis of the activation mechanism of Pseudomonas stutzeri cytochrome c peroxidase through an electron transfer chain”, (2011) Journal of Biological Inorganic Chemistry, 16, 881-888.

4. Cristina G. Timóteo, Alice S. Pereira, Carlos E. Martins, Sunil G. Naik, Américo G. Duarte, José J. G. Moura, Pedro Tavares, Boi Hanh Huynh and Isabel Moura, “Low-Spin Heme b3 in the Catalytic Center of Nitric Oxide Reductase from Pseudomonas nautica”(2011) Biochemistry, 50, 4251-4262.

5. Katja Conrath, Alice S. Pereira, Carlos E. Martins, Cristina G. Timóteo, Pedro Tavares, Silvia Spinelli, Joerg Kinne, Christophe Flaudrops, Christian Cambillau, Serge Muyldermans, Isabel Moura, José J. G. Moura, Mariella Tegoni and Aline Desmyter, “Camelid Nanobodies Raised against an Integral Membrane Enzyme, Nitric Oxide Reductase”, (2009) Protein Science, 18, 619-628.

6. Cristina M. Cordas, Alice S. Pereira, Carlos E. Martins, Cristina G. Timóteo, Isabel Moura, José J. G. Moura and Pedro Tavares, “Nitric Oxide Reductase: Direct Electrochemistry and Electrocatalytic Activity” (2006) ChemBioChem, 7 (12), 1878-81.

7. Cecília Bonifácio, João M. Dias, Carlos Cunha, Axel Muller, Cristina G. Timóteo, Isabel Moura and Maria J. Romão, “Crystallization and Preliminary X-Ray Diffraction Analysis of the Di-heme Cytochrome c Peroxidase from Pseudomonas stutzeri” (2003) Acta Cryst, D59, 345-347.

8. Cristina G. Timóteo, Pedro Tavares, Celia F. Goodhew, Luís C. Duarte, Kornelia Jumel, Francisco M.F. Gírio, Steve Harding, Graham W. Pettigrew and Isabel Moura, “Calcium and Bacterial Peroxidases - The cytochrome c peroxidase from Pseudomonas stutzeri “ (2003) J Biol Inorg Chem, 8, 29-37.