TY - JOUR
T1 - The cadmium binding domains in the metallothionein isoform Cd 7-MT10 from Mytilus galloprovincialis revealed by NMR spectroscopy
AU - Digilio, Giuseppe
AU - Bracco, Chiara
AU - Vergani, Laura
AU - Botta, Mauro
AU - Osella, Domenico
AU - Viarengo, Aldo
PY - 2009/2
Y1 - 2009/2
N2 - The metal-thiolate connectivity of recombinant Cd7-MT10 metallothionein from the sea mussel Mytilus galloprovincialis has been investigated for the first time by means of multinuclear, multidimensional NMR spectroscopy. The internal backbone dynamics of the protein have been assessed by the analysis of 15N T 1 and T 2 relaxation times and steady state {1H}-15N heteronuclear NOEs. The 113Cd NMR spectrum of mussel MT10 shows unique features, with a remarkably wide dispersion (210 ppm) of 113Cd NMR signals. The complete assignment of cysteine Hα and Hβ proton resonances and the analysis of 2D 113Cd-113Cd COSY and 1H- 113Cd HMQC type spectra allowed us to identify a four metal-thiolate cluster (α-domain) and a three metal-thiolate cluster (β-domain), located at the N-terminal and the C-terminal, respectively. With respect to vertebrate MTs, the mussel MT10 displays an inversion of the α and β domains inside the chain, similar to what observed in the echinoderm MT-A. Moreover, unlike the MTs characterized so far, the α-domain of mussel Cd7-MT10 is of the form M4S12 instead of M 4S11, and has a novel topology. The β-domain has a metal-thiolate binding pattern similar to other vertebrate MTs, but it is conformationally more rigid. This feature is quite unusual for MTs, in which the β-domain displays a more disordered conformation than the α-domain. It is concluded that in mussel Cd7-MT10, the spacing of cysteine residues and the plasticity of the protein backbone (due to the high number of glycine residues) increase the adaptability of the protein backbone towards enfolding around the metal-thiolate clusters, resulting in minimal alterations of the ideal tetrahedral geometry around the metal centres.
AB - The metal-thiolate connectivity of recombinant Cd7-MT10 metallothionein from the sea mussel Mytilus galloprovincialis has been investigated for the first time by means of multinuclear, multidimensional NMR spectroscopy. The internal backbone dynamics of the protein have been assessed by the analysis of 15N T 1 and T 2 relaxation times and steady state {1H}-15N heteronuclear NOEs. The 113Cd NMR spectrum of mussel MT10 shows unique features, with a remarkably wide dispersion (210 ppm) of 113Cd NMR signals. The complete assignment of cysteine Hα and Hβ proton resonances and the analysis of 2D 113Cd-113Cd COSY and 1H- 113Cd HMQC type spectra allowed us to identify a four metal-thiolate cluster (α-domain) and a three metal-thiolate cluster (β-domain), located at the N-terminal and the C-terminal, respectively. With respect to vertebrate MTs, the mussel MT10 displays an inversion of the α and β domains inside the chain, similar to what observed in the echinoderm MT-A. Moreover, unlike the MTs characterized so far, the α-domain of mussel Cd7-MT10 is of the form M4S12 instead of M 4S11, and has a novel topology. The β-domain has a metal-thiolate binding pattern similar to other vertebrate MTs, but it is conformationally more rigid. This feature is quite unusual for MTs, in which the β-domain displays a more disordered conformation than the α-domain. It is concluded that in mussel Cd7-MT10, the spacing of cysteine residues and the plasticity of the protein backbone (due to the high number of glycine residues) increase the adaptability of the protein backbone towards enfolding around the metal-thiolate clusters, resulting in minimal alterations of the ideal tetrahedral geometry around the metal centres.
KW - Cadmium-113
KW - Dynamics
KW - Metallothionein
KW - Multinuclear NMR
KW - Protein structure
UR - http://www.scopus.com/inward/record.url?scp=58849146988&partnerID=8YFLogxK
U2 - 10.1007/s00775-008-0435-y
DO - 10.1007/s00775-008-0435-y
M3 - Article
SN - 0949-8257
VL - 14
SP - 167
EP - 178
JO - Journal of Biological Inorganic Chemistry
JF - Journal of Biological Inorganic Chemistry
IS - 2
ER -