8e7f739c6c1329e517592e38a599a650 pmc_28684668.pdf a01cb85ebbe8ecb99de93bab15718f8f79f68e16 pmc_28684668.pdf 166b1f5751bf2416b70c0273975b1d1df49f6093d7765abc60837351f15a7f69 pmc_28684668.pdf Title: Analysis of Metal-Binding Features of the Wild Type and Two Domain-Truncated Mutant Variants of Littorina littorea Metallothionein Reveals Its Cd-Specific Character Subject: After the resolution of the 3D structure of the Cd9-aggregate of the Littorina littorea metallothionein (MT), we report here a detailed analysis of the metal binding capabilities of the wild type MT, LlwtMT, and of two truncated mutants lacking either the N-terminal domain, Lltr2MT, or both the N-terminal domain, plus four extra flanking residues (SSVF), Lltr1MT. The recombinant synthesis and in vitro studies of these three proteins revealed that LlwtMT forms unique M9-LlwtMT complexes with Zn(II) and Cd(II), while yielding a complex mixture of heteronuclear Zn,Cu-LlwtMT species with Cu(I). As expected, the truncated mutants gave rise to unique M6-LltrMT complexes and Zn,Cu-LltrMT mixtures of lower stoichiometry with respect to LlwtMT, with the SSVF fragment having an influence on their metal binding performance. Our results also revealed a major specificity, and therefore a better metal-coordinating performance of the three proteins for Cd(II) than for Zn(II), although the analysis of the Zn(II)/Cd(II) displacement reaction clearly demonstrates a lack of any type of cooperativity in Cd(II) binding. Contrarily, the analysis of their Cu(I) binding abilities revealed that every LlMT domain is prone to build Cu4-aggregates, the whole MT working by modules analogously to, as previously described, certain fungal MTs, like those of C. neoformans and T. mesenterica. It is concluded that the Littorina littorea MT is a Cd-specific protein that (beyond its extended binding capacity through an additional Cd-binding domain) confers to Littorina littorea a particular adaptive advantage in its changeable marine habitat. Keywords: Littorina littorea; metallothionein; metal binding; tridominial MT Author: Òscar Palacios, Elena Jiménez-Martí, Michael Niederwanger, Selene Gil-Moreno, Oliver Zerbe, Sílvia Atrian, Reinhard Dallinger and Mercè Capdevila Creator: LaTeX with hyperref package Producer: pdfTeX-1.40.17 CreationDate: Thu Jul 6 08:09:56 2017 CEST ModDate: Thu Jul 6 08:09:56 2017 CEST Tagged: no UserProperties: no Suspects: no Form: none JavaScript: no Pages: 16 Encrypted: no Page size: 595.276 x 841.89 pts (A4) Page rot: 0 File size: 2451675 bytes Optimized: no PDF version: 1.5 name type encoding emb sub uni object ID ------------------------------------ ----------------- ---------------- --- --- --- --------- CZFEDL+URWPalladioL-Ital Type 1 Custom yes yes no 90 0 PYEPJK+URWPalladioL-Bold Type 1 Custom yes yes no 91 0 CSDNSU+URWPalladioL-BoldItal Type 1 Custom yes yes no 92 0 OIRFHH+VnURWPalladioL-Bold Type 1 Custom yes yes no 93 0 DPRURN+URWPalladioL-Roma Type 1 Custom yes yes no 94 0 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