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The subnanometer resolution structure of the glutamate synthase 1.2-MDa hexamer by cryoelectron microscopy and its oligomerization behavior in solution - Functional implications
被引:23
作者:
Cottevieille, Magali
[2
,3
]
Larquet, Eric
[2
,3
]
Jonic, Slavica
[2
,3
]
Petoukhov, Maxim V.
[4
,5
]
Caprini, Gianluca
[1
]
Paravisi, Stefano
[1
]
Svergun, Dmitri I.
[4
,5
]
Vanoni, Maria A.
[1
]
Boisset, Nicolas
[2
,3
]
机构:
[1] Univ Milan, Dipartimento Sci Biomol & Biotecnol, I-20133 Milan, Italy
[2] Univ Paris 06, Dept Biol Struct, CNRS, IMPMC UMR 7590, F-75252 Paris, France
[3] Univ Paris 07, IPGP, F-75252 Paris, France
[4] DESY, European Mol Biol Lab, Hamburg Outstn, D-22603 Hamburg, Germany
[5] Russian Acad Sci, Inst Crystallog, Moscow 117333, Russia
关键词:
D O I:
10.1074/jbc.M708529200
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 three-dimensional structure of the hexameric (alpha beta)(6) 1.2-MDa complex formed by glutamate synthase has been determined at subnanometric resolution by combining cryoelectron microscopy, small angle x-ray scattering, and molecular modeling, providing for the first time a molecular model of this complex iron-sulfur flavoprotein. In the hexameric species, interprotomeric alpha-alpha and alpha-beta contacts are mediated by the C-terminal domain of the alpha subunit, which is based on a beta helical fold so far unique to glutamate synthases. The alpha beta protomer extracted from the hexameric model is fully consistent with it being the minimal catalytically active form of the enzyme. The structure clarifies the electron transfer pathway from the FAD cofactor on the beta subunit, to the FMN on the alpha subunit, through the low potential [4Fe-4S](1+/2+) centers on the beta subunit and the [3Fe-4S](0/1+) cluster on the alpha subunit. The(alpha beta)(6) hexamer exhibits a concentration-dependent equilibrium with alpha beta monomers and (alpha beta)(2) dimers, in solution, the hexamer being destabilized by high ionic strength and, to a lower extent, by the reaction product NADP(+). Hexamerization seems to decrease the catalytic efficiency of the alpha beta protomer only 3-fold by increasing the K-m values measured for L-Gln and 2-OG. However, it cannot be ruled out that the (alpha beta)(6) hexamer acts as a scaffold for the assembly of multienzymatic complexes of nitrogen metabolism or that it provides a means to regulate the activity of the enzyme through an as yet unknown ligand.
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页码:8237 / 8249
页数:13
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