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Biogenesis of [Fe-S] proteins

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1 Biogenesis of [Fe-S] proteins
Frédéric Barras, LCB, CNRS, Marseille

2 Some [Fe-S] clusters Kiley and Beinert, 2003
Trouver differents types de clusters Kiley and Beinert, 2003

3 iscAnif nifS NIF nifU iscR iscS iscU iscA hscB hscA fdx ISC sufA sufB sufC sufD sufS sufE SUF csdA ygdK CSD

4 Three machineries in E. coli
iscR iscS iscU iscA hscB hscA fdx ISC sufA sufB sufC sufD sufS sufE SUF csdA ygdK CSD

5 Cysteine S donor Fe S acceptor Alanine S Fe-S Scaffold Fe-S Target

6 SUF machinery Cysteine S donor Fe S acceptor Alanine S Fe-S Scaffold ABC ATPase ? Fe-S Target

7 S donor S acceptor Scaffold
ISC machinery Cysteine S donor S acceptor Fe Fe-S Chaperone Alanine Fe-S Scaffold Ferredoxin ? Fe-S Target

8 CSD machinery Cysteine S donor Fe S acceptor Alanine S Fe-S Target

9 Substrate specificity
Suf Isc Iron assimilation Thionucleosides Fe-S Thiamin Molybdopterin Csd

10 Environmental specificity
Suf Isc H2O2 OxyR IscR [2Fe-2S] Fur Csd Nachin et al., 2001,, Schwartz et al., 2001, Zheng et al., 2001, Hantke, 2002 Lee et al., 2004

11 Biogenesis of [Fe-S] proteins in Cenibacterium
Phenotypic complementation E. coli iscS and sufS mutants Library Cenibacterium

12 iscS of Cenibacterium Complementation: thiamine and nicotinic acid requirement Cloned DNA Genome analysis iscR iscS iscU iscA hscB hscA fdx 52 % 69 % 82 % 67 % 39 % 59 % 72 % Genetic organisation identical to E. coli

13 Characterisation of IscSCeni
Purification (His)6-IscSCeni IscSCeni Cysteine desulfurase activity PLP/protein µmol alanine/min-1 mig-1 IscScoli IscSCeni CsdAcoli

14 No SUF machinery in Cenibacterium
Complementation : no sufS-containing insert Genome analysis : one cysteine desulfurase = IscS

15 E. coli sufS mutant complementation
(Growth in presence of iron chelator ) Cloned DNA orf1 cysA Peptidase sodC Carboxynorspermidine dehydrogenase cysW cysT 47% id. Cbl 45 % id. CysB Sulfur metabolism regulator

16

17 One machinery in Cenibacterium
iscR iscS iscU iscA hscB hscA fdx ISC

18 One machinery in Cenibacterium
iscR iscS iscU iscA hscB hscA fdx ISC Implication of the ISC machinery in arsenic resistance Model to study Scaffold proteins Problems to construct mutants……. iscS, iscA, msrA

19 Gabriel Rincón Enríquez
Frédéric Barras Béatrice Py Gabriel Rincón Enríquez Collaborators CEA, Grenoble Sandrine Ollagnier de Choudens Marc Fontecave

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