MULTINUCLEAR NMR PROBES OF BIOLOGICAL SYSTEMS

这个小组的目标围绕着使用复杂的 多核核磁共振技术，以阐明溶液结构，分子 各种生物系统的动力学和作用机制。 在过去的一年里，我们已经扩大了我们的NMR调查的机制， 以及多种金属离子在E. 大肠杆菌碱性酶 磷酸酶。 这些研究包括磷酸酶的3lp NMR 中间体，113 Cd 2离子取代的酶上的113 Cd NMR和13 C和15 N 用（γ-13 C）-和/或（15 N）-生物合成标记的酶的NMR pi）-组氨酸。 用~（113）Cd核磁共振谱测定了化合物的结构。 金属硫蛋白两种主要同工蛋白中的多个金属结合位点 (MT)兔肝、肾和蟹肝胰腺。 兔子MT 发现含有两个独立的金属簇，一个含有4个113 Cd离子 另一个包含3。 螃蟹MT含有两个3-金属簇。 详细 与该蛋白质中的金属离子有关的结构和动力学数据是 这对理解这种蛋白质的生物学功能至关重要。 化学 和13 C NMR技术也已用于确定组成， M和N血型活性唾液酸糖肽的结构方面， 人血型糖蛋白A 这些研究表明，M和N活性 唾液酸糖-八肽都具有相同的寡糖结构。 这些 数据与其中M或N决定因素是 氨基末端氨基酸和NeuNAc残基。 最后，初步研究 正在评估冷冻酶学等技术的潜力， 在常规NMR方法受限的生物系统中的固态NMR 由于化学交换调制。

The objectives of this group revolve around the use of sophisticated multinuclear NMR techniques to elucidate the solution strucutre, molecular dynamics and mechanism of action of a variety of biological systems. During the past year we have extended our NMR investigations of the mechanistic and structural role of the multiple metal ions in the E. coli enzyme alkaline phosphatase. These studies have included 3lp NMR of the enzyme phosphate intermediates, 113Cd NMR on the 113Cd2 ion substituted enzyme and 13C and 15N NMR on the enzyme labeled biosynthetically with (gamma-13C)-and/or (15 N pi)-histidine. 113Cd NMR has been used to determine the structure of the multiple metal-binding sites in the two major isoproteins of metallothionein (MT) from rabbit liver and kidney, and from crab hepatopancreas. Rabbit MT's were found to contain two separate metal clusters, one containing 4 113Cd ions and the other containing 3. Crab MT's contain two 3-metal clusters. Detailed structural and kinetic data pertaining to the metal ions in this protein is essential to understanding the biological function of this protein. Chemical and 13C NMR techniques have also been used to determine the composition and structural aspects of the blood group M and N active sialoglycopeptides from human glycophorin A. These studies have related that the M and N-active sialoglyco-octapeptides both have identical oligosaccharide structures. These data are consistent with a model in which the M or N determinant is the aminoterminal amino acid and a NeuNAc residue(s). Lastly, preliminary studies are underway to evaluate the potential of techniques such as cryoenzymology and solid state NMR in biological systems where conventional NMR methods are limited due to chemical exchange modulation.