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Struct. and funct. study of mR machinery involved in biosynth and insertion of the organoMe+ active site of [FeFe]-H2ase
Transcript of Struct. and funct. study of mR machinery involved in biosynth and insertion of the organoMe+ active site of [FeFe]-H2ase
Institut de Biologie Structurale - Grenoble
Structural and functional study of molecular machinery
involved in biosynthesis and insertion of the organometallic
active site of [FeFe]-hydrogenases
by roman rohac
GTPase activity scaffold protein - insertion of H-cluster
3QQ5, Cendron, L. et al, 2011
1HFE, Fe-only hydrogenase
from Desulfovibrio desulfuricans
Nicolet, Y. et al, 1999
from Thermotoga maritima
3CIX, Nicolet, Y. et al., 2008
uses Tyrosine to produce CO and CN- ligands
[NiFe]-hydrogenases, [Fe]-hydrogenases and
in FeFe-hydrogenases catalytic H-cluster = cubane Fe4S4 cluster
linked to a modified Fe2S2 subcluster with non-proteic ligands
hydrogenases = reversible oxidation of molecular hydrogen
active form structure unknown
insertion mechanism unknown
radical SAM enzymes
catalytic H-cluster from FeFe-hydrogenase
CO & CN- ligands
CO & CN-
X-ray crystallography and biochemistry under reducing conditions
iron-sulfur clusters very sensitive to oxygen
all experiments performed under controlled atmosphere = 95% Nitrogen & 5% Hydrogen
cell culture - harvesting - protein purification - biochemical assays - crystallization - flash freezing - data collection
ligand binding assays and investigation of paramagnetic cluster environement:
liquid state NMR of proteins with paramagnetic FeS clusters
FeS characterization by spectroscopic methods:
M ssbauer, Raman, UV-visible, Electron Paramagnetic Resonance
Hydrogen - clean energy of the future ?
plausibly ones of the very first enzymes
Ph.D. Supervisor: Yvain Nicolet
Ph.D. Director: Juan Carlos Fontecilla-Camps
What's next ?
2 H O
O -tolerant hydrogenase