Determination of the Cd/S Cluster Stoichiometry in Fucus vesiculosus Metallothionein

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• 作者：Maureen E. Merrifield ; Jennifer Chaseley ; Peter Kille ; and Martin J. Stillman
• 刊名：Chemical Research in Toxicology
• 出版年：2006
• 出版时间：March 2006
• 年：2006
• 卷：19
• 期：3
• 页码：365 - 375
• 全文大小：369K
• 年卷期：v.19,no.3(March 2006)
• ISSN：1520-5010

文摘

The seaweed Fucus vesiculosus is unusual when compared with other algal species, in that it cansurvive in toxic-metal-contaminated aquatic environments. The metallothionein gene has been identifiedin F. vesiculosus by Kille and co-workers (Morris, C. A., Nicolaus, B., Sampson, V., Harwood, J. L.,and Kille, P. (1999) Biochem. J. 338, 553), which suggests a possible protective mechanism againsttoxic metals for this species. We report the first detailed study of the metal binding properties of F.vesiculosus metallothionein using UV absorption, circular dichroism (CD), and electrospray mass spectraltechniques. The overall metal-to-sulfur ratios of this novel algal protein when bound to divalent cadmiumand zinc were determined to be Cd₆S₁₆ and Zn₆S₁₆, respectively. Mixed Cd/Zn species were also formedwhen Cd²⁺ was added to the Zn-containing Fucus metallothionein. Only one conformation was identifiedat low pH for the native protein. Analysis of the UV absorption, CD, and ESI-MS spectral data recordedduring stepwise, acid-induced demetalation supports a two-domain structure for the protein, with two3-metal binding sites. The data suggest that one of the domains is significantly less stable than the other,and we tentatively propose from the arrangement of cysteines in the sequence that the two domains areM₃S₇ and M₃S₉ (where M = Cd²⁺ or Zn²⁺). While the M₃S₉ cluster is known in the beta2.gif" BORDER=0 ALIGN="middle"> clusters of crab,lobster, and mammalian metallothioneins, the M₃S₇ is a hitherto unknown cluster structure. Metallothioneinin F. vesiculosus is thought to act as a protective mechanism against incoming toxic metals. The metalbinding studies reported are a putative model for metal binding in vivo.