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.
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.
YhjA 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.
Proposed mode of action of MrpORP in the Fe/S cluster biosynthesis of the ORP complex.
- “Detoxification of hydrogen peroxide by Pathogenic bacteria - E. coli tri-haem peroxidase as a model (PathG_CCP)”, 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.
- “Understanding and Exploiting the Impacts of Low pH on Micro-Organisms (FesBio)” - Cost Action CA18113, Sofia R. Pauleta: National Management Committee Member.
- “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).
Pauleta, SR; Carepo, MSP; Moura, I. 2019. Source and reduction of nitrous oxide. COORDINATION CHEMISTRY REVIEWS, 387, DOI: 10.1016/j.ccr.2019.02.005
Pardoux, R; Fievet, A; Carreira, C; Brochier-Armanet, C; Valette, O; Dermoun, Z; Py, B; Dolla, A; Pauleta, SR; Aubert, C. 2019. The bacterial Mrp(ORP) is a novel Mrp/NBP35 protein involved in iron-sulfur biogenesis. Scientific Reports, 9, DOI: 10.1038/s41598-018-37021-8
Carreira, C; Mestre, O; Nunes, RF; Moura, I; Pauleta, SR. 2018. Genomic organization, gene expression and activity profile of Marinobacter hydrocarbonoclasticus denitrification enzymes. PeerJ, 6, DOI: 10.7717/peerj.5603
Nobrega, CS; Pauleta, SR. 2018. Interaction between Neisseria gonorrhoeae bacterial peroxidase and its electron donor, the lipid-modified azurin. FEBS LETTERS, 592, DOI: 10.1002/1873-3468.13053
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