Tungsten-containing formats dehydrogenase from Desulfovibrio gigas:: metal identification and preliminary structural data by multi-wavelength crystallography

被引:38
作者
Raaijmakers, H [1 ]
Teixeira, S [1 ]
Dias, JM [1 ]
Almendra, MJ [1 ]
Brondino, CD [1 ]
Moura, I [1 ]
Moura, JJG [1 ]
Romao, MJ [1 ]
机构
[1] Univ Nova Lisboa, Fac Ciencias & Tecnol, Ctr Quim Fina & Biotecnol, Dept Quim, P-2825114 Caparica, Portugal
来源
JOURNAL OF BIOLOGICAL INORGANIC CHEMISTRY | 2001年 / 6卷 / 04期
关键词
formate dehydrogenase; tungsten-containing enzymes; molybdopterin; iron-sulfur centers; multi-wavelength anomalous dispersion;
D O I
10.1007/s007750100215
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 tungsten-containing formate dehydrogenase (W-FDH) isolated from Desulfovibrio gigas has been crystallized in space group P2(1), with cell parameters a=73.8 Angstrom, b=111.3 Angstrom, c=156.6 Angstrom and beta =93.7 degrees. These crystals diffract to beyond 2.0 Angstrom on a synchrotron radiation source. W-FDH is a heterodimer (92 kDa and 29 kDa subunits) and two W-FDH molecules are present in the asymmetric unit. Although a molecular replacement solution was found using the periplasmic nitrate reductase as a search model, additional phasing information was needed. A multiple-wavelength anomalous dispersion (MAD) dataset was collected at the W- and Fe-edges, at four different wavelengths. Anomalous and dispersive difference data allowed us to unambiguously identify the metal atoms bound to W-FDH as one W atom with a Se-cysteine ligand as well as one [4Fe-4S] cluster in the 92 kDa subunit, and three additional [4Fe-4S] centers in the smaller 29 kDa subunit. The D. gigas W-FDH was previously characterized based on metal analysis and spectroscopic data. One W atom was predicted to be bound to two molybdopterin guanine dinucleotide (MGD) pterin cofactors and two [4Fe-4S] centers were proposed to be present. The crystallographic data now reported reveal a selenium atom (as a Se-cysteine) coordinating to the W site, as well as two extra [4Fe-4S] clusters not anticipated before. The EPR data were re-evaluated in the Light of these new results.
引用
收藏
页码:398 / 404
页数:7
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