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Sulfur trafficking in Escherichia coli: The role of CsdL protein

Master in Chemistry - Sujet de stage de Master 2

Published on 18 September 2017

Laboratory: Chemistry and Biology of Metals laboratory
Head: Dr. Stéphane Ménage
Name of the team: Biocatalyse (BioCat)
Head of the team: Dr. Sandrine Ollagnier de Choudens
Name and quality of the internship’s head: Dr. Sandrine Ollagnier de Choudens, PhD, Dr2 CNRS, HDR
Address: Bâtiment K’, 17 avenue des martyrs, 38054 Grenoble cedex 9
Phone: 04 38 78 91 22

Sulfur trafficking in Escherichia coli: The role of CsdL protein

Internship's main goals:
Understand at a molecular level the role of CsdL protein in the sulfur trafficking in Escherichia coli. In particular, we aim at understand how CsdL, a protein endowed with a sulfur transfer activity, is involved in the formation of a post-translational modification of a tRNA that does not contain any sulfur atom. The project will investigate the hypothesis of a possible cryptic mechanism in this tRNA modification.

Educational interests and skills targeted:
The project combines multidisciplinary approaches such as chemistry (synthesis of the substrate), biochemistry (proteins purification, enzymatic assays) and biophysics (MS).

Sulfur adds considerable functionality to a wide variety of biomolecules because of its unique properties: its chemical bonds are made as well as broken easily and sulfur equally well serves as an electrophile (e.g. in disulfides) and as a nucleophile (e.g. as thiol). For incorporation into biomolecules, sulfur must be reduced and/or activated, and sulfate or polysulfides are substrates for reductases in nature. With sulfide as the final product of reduction, incorporation into cysteine is possible and makes this amino acid a central building block for many sulfur compounds. Sulfur is mobilized from L-cysteine through PLP-dependent enzymes that are called cysteine desulfurases. Their activity allows the formation of organic sulfur molecules and Fe / S inorganic clusters. Escherichia coli contains three cysteine desulfurases, named ISCS, SufS and CsdA. We have shown in the laboratory that CsdA interacts with two other proteins, CsdE and CsdL which take part of the csdA-csdE-csdL operon. It was shown on the laboratory that CsdE stimulates the activity of CsdA but the function of CsdL is not known yet although we know that CsdL receives sulfur atoms from CsdA and/or CsdE through transpersulfuration reactions. CsdA, CsdE and CsdL have been shown to be involved in vivo in post-translational modification of a tRNA which curiously does not contain sulfur. The proposed project aims at understanding the mechanism at a molecular level of this modification involving CsdA, CsdE and CsdL. Our hypothesis, that will be checked, is that the mechanism involves an activation by sulfur which is then removed (sulfur cryptic mechanism).

Approaches & material used:
Chemistry (synthesis of the substrate), biochemistry (proteins purification, enzymatic assays) and biophysics. Materials used: HPLC, FPLC, spectrophotometers, mass spectrometry and glove boxes.

Technical skills needed:
A biochemist or chemist with good notions in bioinorganic chemistry.

Internship’s date:
January to june 2018
In order to meet the deadline for entry to the CEA (3 months for the SPAS), the candidate must present himself as soon as possible.

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