Armitage I M, Dalgarno D C, Johnson B A
Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, CT 06510.
Experientia Suppl. 1987;52:159-69. doi: 10.1007/978-3-0348-6784-9_9.
Multinuclear 1 and 2 dimensional magnetic resonance methods have been used to investigate the structures and metal binding properties of metallothioneins (MTs) isolated from several different sources. 113Cd NMR studies have unambiguously shown that the 7 g-atoms of Cd2+ bound per mole of the mammalian MT are located in two separate metal clusters, one containing 4 metal ions and the other, 3 metal ions. In the invertebrate (Scylla serrata) MT, similar studies have revealed that the 6 g-atoms of bound Cd2+ are distributed in two distinct 3-metal clusters while in Neurospora MT, the 3 g-atoms of bound Cd2+ are arranged in a pseudo 3-metal cluster. With the exception of one of the Cd2+ sites in this latter cluster, all the Cd2+ ions are tetrahedrally coordinated to four cysteine thiolate ligands with single cysteinyl sulfurs bridging adjacent metals. These conclusions are based on the 113Cd chemical shift data and a detailed analysis of the observed 113Cd-113Cd scalar couplings by both homonuclear decoupling and 2D techniques. In addition, the 113Cd NMR studies have revealed significant differences in the affinity of different metal ions for the two mammalian metal clusters. For the 3-metal cluster, the affinity is found to decrease in the order Cu+ greater than Cd2+ greater than Zn2+ with Cd2+ greater than Zn2+ for the 4 metal cluster and Cd2+ (4-metal cluster) greater than Cd2+ (3-metal cluster). The 113Cd NMR data are currently being integrated with 500 MHz 2D 1H and 1H-113Cd chemical shift correlated multiple quantum data sets to more completely define the structural arrangement of the metal clusters in the tertiary structure of these proteins.
多核一维及二维磁共振方法已被用于研究从几种不同来源分离得到的金属硫蛋白（MTs）的结构和金属结合特性。113Cd NMR研究明确表明，每摩尔哺乳动物MT结合的7个克原子Cd2+位于两个独立的金属簇中，一个包含4个金属离子，另一个包含3个金属离子。在无脊椎动物（锯缘青蟹）MT中，类似研究表明，结合的6个克原子Cd2+分布在两个不同的三金属簇中，而在粗糙脉孢菌MT中，结合的3个克原子Cd2+排列成一个假三金属簇。除了后一个簇中的一个Cd2+位点外，所有Cd2+离子都通过单个半胱氨酸硫原子桥连相邻金属，与四个半胱氨酸硫醇盐配体形成四面体配位。这些结论基于113Cd化学位移数据以及通过同核去耦和二维技术对观察到的113Cd-113Cd标量耦合的详细分析。此外，113Cd NMR研究还揭示了不同金属离子对两种哺乳动物金属簇亲和力的显著差异。对于三金属簇，发现亲和力按Cu+＞Cd2+＞Zn2+的顺序降低，对于四金属簇为Cd2+＞Zn2+，且Cd2+（四金属簇）＞Cd2+（三金属簇）。目前，113Cd NMR数据正在与500 MHz二维1H和1H-113Cd化学位移相关多量子数据集相结合，以更全面地确定这些蛋白质三级结构中金属簇的结构排列。
