Research Lab

Microbial Stress

 Microbial Stress
29676

Lab Members

Lab leader

Research Interests

Our projects focus on the characterization of molecular systems (mainly metalloproteins) that are responsible for metal tolerance in bacteria, involved in the detoxification of reactive oxygen species from pathogenic and non-pathogenic bacteria, and an ancient iron-sulfur cluster biosynthesis/ repair from strict anaerobic bacteria. For that we use several spectroscopic (visible, EPR, NMR and resonance Raman), and biophysical (steady-state and fast-kinetics, microcalorimetry and analytical ultracentrifugation) techniques, complemented by proteomics and transcriptomic analysis. Biomolecular NMR is also used to structurally characterize these (metallo)proteins and protein interactions.

Research Highlights
Bacterial Peroxidases from Neisseria gonorrhoeae and Escherichia coli

Bacterial Peroxidases are periplasmic enzymes involved in the reduction of hydrogen peroxide in the periplasm of pathogenic bacteria, and are considered to be a first line defence mechanism against hydrogen peroxide during infection. The di-heme bacterial peroxidase from N. gonorrhoeae was spectroscopic and biochemically characterized, and shown to receive electrons from the lipid modified azurin from the same organism. The tri-heme bacterial peroxidase from E. coli was isolated and biochemically characterized for the first time. This enzyme is a quinol peroxidase that is expressed under anaerobic conditions.

 

fig1YhjA role in E. coli when the cells are transiently exposed to oxygen.

 

The ORP Complex is involved in the biosynthesis of Fe/S clusters in strict anaerobic bacteria

One of the proteins that form the ORP complex from Desulfovibrio alaskensis G20 was isolated and identified as an Fe-S protein. This protein was spectroscopically characterized as binding one or two [2Fe-2S] clusters. Other proteins involved in this protein complex were also isolated for the first time and identified to belong to the Nbp35 family. These proteins were shown to donate Fe/S clusters to target apo-proteins in vitro, and thus might complement the minimal Fe/S biosynthesis system found in sulfate reducing bacteria.

 

fig2Proposed mode of action of MrpORP in the Fe/S cluster biosynthesis of the ORP complex.

 

Representative Projects

  • “Hydrogen Peroxide Detoxification by Pathogenic Bacteria - E. coli Three-hemic peroxidase as a model”, FCT-MCTES, Total and Unit funding: € 237,634, Sofia R. Pauleta (PI).
  • “The Biogenesis of Iron-sulfur Proteins: from Cellular Biology to Molecular Aspects (FeSBioNet)” - Cost Action CA15133, Sofia R. Pauleta: substitute MC & Communication´s Representative.
  • “Functional and structural characterization of unknown function conserved metalloproteins in anaerobes: a putative role in the control of cell division?”, FCT-MCTES, Total and Unit funding: € 225.280.00, Sofia R. Pauleta (PI).
  • “The metalloproteins involved in cell division - Functional and Structural Characterization”, Pessoa 2014 between REQUIMTE and CNRS, Total funding: €2,000 Sofia R. Pauleta (PI).
  • “Insights into novel bacterial cytochrome c peroxidases from pathogenic bacteria, Neisseria gonorrhoeae and Escherichia coli”, FCT-MCTES, Total and Unit funding: € 191,979, Sofia R. Pauleta (PI).

 

Selected Publications

Nobrega, CS; Devreese, B; Pauleta, SR. 2018. YhjA - An Escherichia coli trihemic enzyme with quinol peroxidase activity. BIOCHIMICA ET BIOPHYSICA ACTA-BIOENERGETICS, 1859, DOI: 10.1016/j.bbabio.2018.03.008
Cíntia Carreira; Sofia RPauleta; Isabel Moura. 2017. The catalytic cycle of nitrous oxide reductase — The enzyme that catalyzes the last step of denitrification. JOURNAL OF INORGANIC BIOCHEMISTRY, DOI: 10.1016/j.jinorgbio.2017.09.007
Susana Ramos; Rui MAlmeida; Cristina MCordas; José JGMoura; Sofia RPauleta; Isabel Moura. 2017. Insights into the recognition and electron transfer steps in nitric oxide reductase from Marinobacter hydrocarbonoclasticus. JOURNAL OF INORGANIC BIOCHEMISTRY, DOI: 10.1016/j.jinorgbio.2017.09.001
Johnston, EM; Carreira, C; Dell'Acqua, S; Dey, SG; Pauleta, SR; Moura, I; Solomon, EI. 2017. Spectroscopic Definition of the Cu-Z degrees Intermediate in Turnover of Nitrous Oxide Reductase and Molecular Insight into the Catalytic Mechanism. JOURNAL OF THE AMERICAN CHEMICAL SOCIETY, 139, DOI: 10.1021/jacs.6b13225
Carreira, C; Carepo, MSP; Todorovic, S; Aubert, C; Pauleta, SR. 2017. The Orange Protein Complex - Novel Fe-S proteins from sulfate reducing bacteria. JOURNAL OF BIOLOGICAL INORGANIC CHEMISTRY, 22, DOI: