Biogenesis of [Fe-S] proteins in Escherichia coli

1
Frederic Barras, LCB, CNRS, Marseille
Biogenesis of Fe-S proteins in Escherichia coli
Marburg, 21 april 2004
2
Biogenesis of Fe-S proteins in Escherichia coli

• Background
• The IscS system others story
• The Suf system our story
• The Csd system the new story

3
Some Fe-S clusters
Kiley and Beinert, 2003
4
Biological functions of Fe-S clusters

• Electron transfer
• Sulfur transfer
• Sensing O2 and derivatives

5
Fe-S cluster containing proteins FNR
PJ Kiley
6
Fe-S cluster containing proteins Aconitase/IRE-B
P
Aconitase 4Fe-4S cluster IRE-binding site not
accessible
IRE-BP No 4Fe-4S cluster IRE-binding site
accessible
7
Fe-S cluster containing proteins SoxR
2Fe-2S2 2Fe-2S2
-35
-10
2Fe-2S3
2Fe-2S3
-35
-10
soxS mRNA
Pomposiello and Demple, 2001
8
How to make Fe-S proteins? 1- Chemists answer
Apo-protein
Fe-S protein
Fe2 Na2S DTT
9
How to make Fe-S proteins? 2- Biologists answer

• Source of Fe and S?
• (toxicity)
• Assembly of Fe-S cluster
• (insertion and folding)

10
Biogenesis of Fe-S proteins in Escherichia coli

• Background
• The IscS system others story
• The Suf system our story
• The Csd system the new story

11
Study of nitrogen fixation in Azotobacter
vinelandii Dr. Deans laboratory (1993)
Apo-nitrogenase
4Fe-4S nitrogenase
Fe2 Cysteine DTT NifS
12
Enzymatic activity of Cysteine desulfurases
13
NifS-like gene clusters
Frazzon and Dean, 2003 Muhlenhoff and Lill, 2000
14
Functions of the isc gene products
iscR
iscS
iscU
iscA
hscB
hscA
fdx
-
Cysteine desulfurase
2Fe-2S Ferredoxin
2Fe-2S
Fe-S cluster assembly scafold
H2O2
Molecular chaperones
15
Model
ADP
HscA
ATP
B
Apo
IscU
Cysteine
Fe-S
IscS
S
Alanine
Cysteine
IscS
S
IscA
Fe-S
Fe-S
Fdx
Alanine
16
3 NifS-like in E. coli
iscS
csdA (CSD)
sufS (csdB)
location
53.7
37.8
63.4
Cysteine Selenocysteine
0.38 3.1
0.9 6.2
0.02 5.5
I
II
II
AminotransferaseClass V
SSGSACTS
RXGHHCA


Structure 3D
17
Loci containing NifS homologues in E. coli
iscR
iscS
iscU
iscA
sufC
sufS
sufE
sufA
sufB
sufD
csdA
ygdK
ygdL
18
Biogenesis of Fe-S proteins in Escherichia coli

• Background
• The IscS system others story
• The Suf system our story
• The Csd system the new story

19
Erwinia chrysanthemi
Enterobacteria Plant pathogen Search for
virulence genes by transposon mutagenesis
Beaulieu and van Gijsegem, J Bact, 1990
20
pin10 suf genes
RT-PCR C PCR
sufC
sufD
sufS
sufE
sufA
sufB

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