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Published December 2010 | Supplemental Material
Journal Article Open

Structural insights into the enzyme catalysis from comparison of three forms of dissimilatory sulphite reductase from Desulfovibrio gigas

Abstract

The crystal structures of two active forms of dissimilatory sulphite reductase (Dsr) from Desulfovibrio gigas, Dsr-I and Dsr-II, are compared at 1.76 and 2.05 Å resolution respectively. The dimeric α_2β_2γ_2 structure of Dsr-I contains eight [4Fe–4S] clusters, two saddle-shaped sirohaems and two flat sirohydrochlorins. In Dsr-II, the [4Fe–4S] cluster associated with the sirohaem in Dsr-I is replaced by a [3Fe–4S] cluster. Electron paramagnetic resonance (EPR) of the active Dsr-I and Dsr-II confirm the co-factor structures, whereas EPR of a third but inactive form, Dsr-III, suggests that the sirohaem has been demetallated in addition to its associated [4Fe–4S] cluster replaced by a [3Fe–4S] centre. In Dsr-I and Dsr-II, the sirohydrochlorin is located in a putative substrate channel connected to the sirohaem. The γ-subunit C-terminus is inserted into a positively charged channel formed between the α- and β-subunits, with its conserved terminal Cysγ104 side-chain covalently linked to the CHA atom of the sirohaem in Dsr-I. In Dsr-II, the thioether bond is broken, and the Cysγ104 side-chain moves closer to the bound sulphite at the sirohaem pocket. These different forms of Dsr offer structural insights into a mechanism of sulphite reduction that can lead to S_3O_6^(2−), S_2O_3^(2−) and S^(2−).

Additional Information

© 2010 Blackwell Publishing Ltd. Accepted 1 September, 2010. Article first published online: 28 Sep. 2010. We are indebted to the Yuch-Cheng Jean and supporting staffs at beamlines BL13B1 and BL13C1 at the National Synchrotron Radiation Research Center (NSRRC), and Jeyakanthan Jeyaraman, Masato Yoshimura and Hirofumi Ishii at the Taiwan contracted beamline BL12B2 at SPring-8, for their assistance. We are also grateful to Go Ueno and his staff for assistance at BL26-I and -II of SPring-8 at RIKEN. Portions of this research were carried out at the NSRRCNCKU Protein Crystallography Laboratory at National Cheng Kung University (NCKU). We thank Yi-Hong Lin for protein sequencing; Yu-Jhang Lu for assistance with the low temperature EPR experiments on the anaerobically purified Dsr proteins; and Wen-Hsiung Li, James Ketudat Cairns and Chien-Te Chen for discussions. This work was supported in part by grants from the National Synchrotron Radiation Research Center (NSRRC) (NSRRC 954RSB03, 963RSB03 and 973RSB02 and from the National Science Council (NSC) (NSC 95-2311-B213-001-MY3, NSC 95-2923-B-213-001- MY3 and NSC 98-2311-B-213-MY3) of Taiwan.

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