The Structure of the Active Site H-Cluster of [FeFe] Hydrogenase from the Green Alga Chlamydomonas reinhardtii Studied by X-ray Absorption Spectroscopy

被引:60
作者
Stripp, Sven [1 ]
Sanganas, Oliver [2 ]
Happe, Thomas [1 ]
Haumann, Michael [2 ]
机构
[1] Ruhr Univ Bochum, Lehrstuhl Biochem Pflanzen, AG Photobiotechnol, D-44780 Bochum, Germany
[2] Free Univ Berlin, Inst Expt Phys, D-14195 Berlin, Germany
关键词
IRON-SULFUR CLUSTERS; DENSITY-FUNCTIONAL THEORY; BIOLOGICAL METAL CENTERS; ONLY HYDROGENASE; ELECTRONIC-STRUCTURE; DESULFOVIBRIO-DESULFURICANS; MANGANESE COMPLEX; LIGHT SENSITIVITY; CLOSTRIDIUM-ACETOBUTYLICUM; CRYSTAL-STRUCTURE;
D O I
10.1021/bi900010b
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
The [FeFe] hydrogenase (CrHydA1) of the green alga Chlamydomonas reinhardtii is the smallest hydrogenase known and can be considered as a "minimal unit" for biological H-2 production. Due to the absence of additional FeS clusters as found in bacterial [FeFe] hydrogenases, it was possible to specifically study its catalytic iron-sulfur cluster (H-cluster) by X-ray absorption spectroscopy (XAS) at the Fe K-edge. The XAS analysis revealed that the CrHydA1 H-cluster consists of a [4Fe4S] cluster and a diiron site, 2Fe(H), which both are similar to their crystallographically characterized bacterial counterparts. Determination of the individual Fe-Fe distances in the [4Fe4S] cluster (similar to 2.7 angstrom) and in the 2FeH unit (similar to 2.5 angstrom) was achieved. Fe-C (=O/N) and Fe-S bond lengths were in good agreement with crystallographic data on bacterial enzymes. The loss of Fe-Fe distances in protein purified under mildly oxidizing conditions indicated partial degradation of the H-cluster. Bond length alterations detected after incubation of CrHydA1 with CO and H-2 were related to structural and oxidation state changes at the catalytic Fe atoms, e.g., to the binding of an exogenous CO at 2Fe(H) in CO-inhibited enzyme. Our XAS studies pave the way for the monitoring of atomic level structural changes at the H-cluster during H-2 catalysis.
引用
收藏
页码:5042 / 5049
页数:8
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