TITLE

Characterization and Crystallization of an IscU-type Scaffold Protein with Bound [2Fe—2S] Cluster from the Hyperthermophile, Aquifex aeolicus

AUTHOR(S)
Shimomura, Yoshimitsu; Kamikubo, Hironari; Nishi, Yoshinori; Masako, Takuya; Kataoka, Mikio; Kobayashi, Yuji; Fukuyama, Keiichi; Takahashi, Yasuhiro
PUB. DATE
November 2007
SOURCE
Journal of Biochemistry;Nov2007, Vol. 142 Issue 5, p577
SOURCE TYPE
Academic Journal
DOC. TYPE
Article
ABSTRACT
IscU plays a key role during iron–sulphur (Fe–S) cluster biosynthesis as a scaffold for the assembly of a nascent, highly labile Fe–S cluster. Here we report the characterization of an IscU-type protein (Aa IscU) from the hyperthermophilic bacterium Aquifex aeolicus. Unlike other known homologues of IscU, expression of Aa IscU in Escherichia coli has yielded an Fe–S cluster-containing holo-protein. Biochemical and spectroscopic studies of the wild-type Aa IscU and its Asp38-to-Ala substituted (D38A) variant molecule indicate that the holo-protein forms a trimer containing substoichiometric [2Fe–2S] cluster with its stability substantially increased by a D38A substitution. The [2Fe–2S] cluster was oxygen-labile and upon loss of the cluster, the resultant apo-form dissociated into a smaller species, a mixture of monomer and dimer with the dimer form predominating. Reddish-brown crystals of holo-Aa IscU-D38A were obtained under anaerobic conditions, that gave diffractions beyond 2.0 Å resolution with synchrotron radiation. The crystal belongs to the space group P21212 with unit-cell parameters a = 72.6, b = 122.3, c = 62.4 Å, where the asymmetric unit contains three molecules of Aa IscU. Successful crystallization of holo-Aa IscU-D38A strongly suggests that the trimer association carrying substoichiometric [2Fe–2S] cluster represents a conformationally stable oligomeric state.
ACCESSION #
44544048

 

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