Haimovich A, Goldbourt A
School of Chemistry, Raymond and Beverly Sackler Faculty of Exact Sciences, Tel Aviv University, Ramat Aviv 69978, Tel Aviv, Israel.
School of Chemistry, Raymond and Beverly Sackler Faculty of Exact Sciences, Tel Aviv University, Ramat Aviv 69978, Tel Aviv, Israel.
J Magn Reson. 2015 May;254:131-8. doi: 10.1016/j.jmr.2015.02.003. Epub 2015 Feb 14.
Lithium, in the form of lithium carbonate, is one of the most common drugs for bipolar disorder. Lithium is also considered to have an effect on many other cellular processes hence it possesses additional therapeutic as well as side effects. In order to quantitatively characterize the binding mode of lithium, it is required to identify the interacting species and measure their distances from the metal center. Here we use magic-angle spinning (MAS) solid-state NMR to study the binding site of lithium in complex with glycine and water (LiGlyW). Such a compound is a good enzyme mimetic since lithium is four-coordinated to one water molecule and three carboxylic groups. Distance measurements to carbons are performed using a 2D transferred echo double resonance (TEDOR) MAS solid-state NMR experiment, and water binding is probed by heteronuclear high-resolution proton-lithium and proton-carbon correlation (wPMLG-HETCOR) experiments. Both HETCOR experiments separate the main complex from impurities and non-specifically bound lithium species, demonstrating the sensitivity of the method to probe the species in the binding site. Optimizations of the TEDOR pulse scheme in the case of a quadrupolar nucleus with a small quadrupole coupling constant show that it is most efficient when pulses are positioned on the spin-1/2 (carbon-13) nucleus. Since the intensity of the TEDOR signal is not normalized, careful data analysis that considers both intensity and dipolar oscillations has to be performed. Nevertheless we show that accurate distances can be extracted for both carbons of the bound glycine and that these distances are consistent with the X-ray data and with lithium in a tetrahedral environment. The lithium environment in the complex is very similar to the binding site in inositol monophosphatase, an enzyme associated with bipolar disorder and the putative target for lithium therapy. A 2D TEDOR experiment applied to the bacterial SuhB gene product of this enzyme was designed to probe direct correlations between lithium, the enzyme inhibitor, and the closest carboxyl carbons of the binding site. At this point, the chemical shift of the bound carboxyl groups in this 29 kDa enzyme could be determined.
碳酸锂形式的锂是治疗双相情感障碍最常用的药物之一。锂还被认为对许多其他细胞过程有影响，因此它具有额外的治疗作用以及副作用。为了定量表征锂的结合模式，需要识别相互作用的物种并测量它们与金属中心的距离。在这里，我们使用魔角旋转（MAS）固态核磁共振来研究锂与甘氨酸和水形成的配合物（LiGlyW）中的锂结合位点。这样的化合物是一种很好的酶模拟物，因为锂与一个水分子和三个羧基形成四配位。使用二维转移回波双共振（TEDOR）MAS固态核磁共振实验对碳进行距离测量，并通过异核高分辨率质子 - 锂和质子 - 碳相关（wPMLG - HETCOR）实验探测水的结合情况。这两个HETCOR实验将主要配合物与杂质和非特异性结合的锂物种区分开来，证明了该方法探测结合位点中物种的灵敏度。对于具有小四极耦合常数的四极核，优化TEDOR脉冲序列表明，当脉冲位于自旋 - 1/2（碳 - 13）核上时效率最高。由于TEDOR信号的强度未归一化，因此必须进行同时考虑强度和偶极振荡的仔细数据分析。尽管如此，我们表明可以提取结合甘氨酸两个碳的准确距离，并且这些距离与X射线数据以及锂在四面体环境中的情况一致。配合物中的锂环境与肌醇单磷酸酶中的结合位点非常相似，肌醇单磷酸酶是一种与双相情感障碍相关的酶，也是锂治疗的假定靶点。应用于该酶的细菌SuhB基因产物的二维TEDOR实验旨在探测锂、酶抑制剂与结合位点中最接近的羧基碳之间的直接相关性。此时，可以确定这种29 kDa酶中结合羧基的化学位移。
