New Nickel Environment for Biology: Cofactor Assembly and Function

该奖项由密歇根州立大学生命过程化学项目资助密歇根州立大学的Robert Hausinger博士和Jian Hu博士研究微生物如何相互转化乳酸的两种镜像结构，乳酸是碳水化合物代谢的中心元素。这种被称为乳酸消旋酶的酶需要蛋白质成分和镍。镍在生物学中很少见。更值得注意的是，乳酸消旋酶中的镍存在于一个以前从未观察到的独特环境中。这个项目调查细菌如何制造创造独特环境的辅因子。这项工作为博士后研究人员、研究生和本科生提供结构和生化机制领域的培训。这些研究人员还参与了与这一主题相关的公众实践教育活动，这些活动将在大学一年一度的科学节上吸引所有年龄段的终身学习者。这项研究阐明了乳酸消旋酶活性所需的四种蛋白质的结构-功能方面。它描述了LLAB如何催化一种新的反应，即烟酸腺嘌呤二核苷酸的吡啶环的羧化，同时水解磷酸酐键生成吡啶-3，5-二羧酸单核苷酸。对LARE(一种牺牲的硫转移酶)的进一步研究，确定了这两个羧酸基是如何转化为硫代羧酸基的，并确定了由此产生的酶的脱氢丙氨酸侧链是否可以再生为半胱氨酸。这项工作揭示了LARC如何催化依赖CTP的镍安装到吡啶-3-5-二硫代甲酸单核苷酸中，这是第一个环金属反应的生物例子，以产生辅因子。最后，这些研究显示了辅因子是如何与不同的LARA物种结合的，并确定了该酶催化的其他潜在转化。结合起来，可以描述这种新型的含镍辅因子的生物合成和作用。这项创新的研究扩展了我们对镍生物化学的理解，并建立了涉及系留镍钳子辅因的氧化还原化学的新范式。这一奖项反映了NSF的法定使命，并通过使用基金会的智力优势和更广泛的影响审查标准进行评估，被认为值得支持。

With this award, the Chemistry of Life Processes Program in the Chemistry Division is funding Dr. Robert Hausinger and Dr. Jian Hu from Michigan State University to characterize how a microorganism interconverts the two mirror image structures of lactic acid, a central element of carbohydrate metabolism. The enzyme that does this, called lactate racemase, requires both a protein component and nickel. Nickel is very rarely found in biology. Even more remarkably, the nickel in lactate racemase is present in a unique environment never before observed. This project investigates how the bacterium makes the cofactor that creates the unique environment. This effort provides training to a postdoctoral researcher, a graduate student, and undergraduate students in the areas of structure and biochemical mechanisms. These researchers also participate in hands-on educational activities for the public related to this topic that will engage lifelong learners of all ages at the annual Science Festival at the university.This research elucidates structure-function aspects of the four proteins required for lactate racemase activity. It characterizes how LarB catalyzes a new reaction, carboxylation of the pyridinium ring of nicotinic acid adenine dinucleotide while hydrolyzing the phosphoanhydride linkage to produce pyridinium-3,5-dicarboxylic acid mononucleotide. Additional studies of LarE, a sacrificial sulfur transferase, establish how the two carboxylic acid groups are transformed into thiocarboxylate groups and define whether the resulting dehydroalanine side chain of the enzyme can be regenerated to cysteine. This work uncovers how LarC catalyzes the CTP-dependent installation of Ni into pyridinium-3-5-dithiocarboxylic acid mononucleotide, the first biological example of a cyclometallation reaction, to create the cofactor. Finally, these investigations show how the cofactor is bound to different LarA species and identifies other potential transformations catalyzed by the enzyme. In combination, the biosynthesis and roles of this novel nickel-containing cofactor can be described. This innovative research expands our understanding of nickel biochemistry and establishes a new paradigm for redox chemistry involving a tethered nickel-pincer cofactor.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.