2-Deoxyglucose Kills Myxococcus Xanthus

黄原粘球菌是一种掠夺性革兰氏阴性细菌，它的代谢能量主要来自氨基酸的氧化。这个物种是黏菌中最具特征的代表，黏菌是唯一一组通过经历多细胞发育程序来对饥饿（蛋白质）做出反应的细菌。以前的研究表明M. xanthus没有“甜食”。也就是说，包括葡萄糖在内的简单己糖不会刺激其生长，即使是在特定的培养基上。然而，黄原草对葡萄糖类似物2-脱氧葡萄糖（2dGIc）敏感。这种抗生素是有毒的，因为它被己糖激酶磷酸化，这是大多数利用葡萄糖的生物体中糖分解代谢的初级糖酵解途径中的第一种酶。抗2dGIc自发突变体的分离和鉴定表明，黄原菌产生己糖激酶，必须运输单糖。这种己糖激酶是不寻常的，因为它被两种必需的己糖糖氨基葡萄糖和n -乙酰氨基葡萄糖所抑制。另一种细菌己糖激酶，由E. coil产生，也被同样的糖所抑制。本研究的三个主要目标是：(1)了解为什么这些酶对2dGIc敏感并被己糖胺抑制；(2)确定大多数生物体内导致2dGIc毒性的2dGIc-6-磷酸的精确靶点；(3)了解这种生物对氨基酸的偏好高于葡萄糖的调节机制。为了完成第一个任务，己糖激酶结合底物和利用这些糖的动力学将使用动力学和机械方法进行研究。本研究将对黄原菌己糖激酶基因进行克隆和测序，并对黄原菌和黄原菌的酶进行纯化。其次，最近发现细胞内2dgic -6-磷酸的产生会导致M. xanthus中ATP的消耗，就像在哺乳动物细胞中一样，这导致了对参与中心碳代谢的关键酶的研究，这种酶被低浓度的2dgic -6-磷酸特异性抑制，再次使用传统的生化策略。第三，将比较容易氧化葡萄糖的M. virescens和不容易氧化葡萄糖的M. xanthus的葡萄糖分解代谢。许多生物，如人类，燃烧糖来获取能量，但更喜欢富含蛋白质的饮食。土壤细菌，黄粘球菌，选择富含蛋白质的饮食（“牛排”）而不是富含碳水化合物的饮食（“土豆”），因为糖不会刺激它的生长。这项研究的目的是了解为什么一些个体生物有这种饮食偏好。生化研究表明，喜欢吃牛排的细菌M. xanthus仍然能尝出糖的味道，因为它被2-脱氧葡萄糖的甜味毒素愚弄了，这种甜味葡萄糖尝起来像富含甜味葡萄糖的佳得乐TM，但效果相反。这种毒素，2-脱氧葡萄糖，50年来一直被认为对大多数生物有毒。没有人真正理解其中的原因，除了在许多活细胞中发现它消耗ATP（能量）水平，包括M. xanthus。M. xanthus将被用作一种模式生物来弄清楚为什么这种毒药是一种毒药，同时，在基本的化学水平上获得关于生物如何选择食物的基本见解。

YouderianThe predatory Gram-negative bacterium, Myxococcus xanthus, derives its metabolic energy primarily from the oxidation of amino acids. This species is the best-characterized representative of the Myxobacteria, the only group of bacteria that respond to starvation (for proteins) by undergoing a program of multicellular development. Previous studies suggested that M. xanthus does not have a "sweet tooth". That is, simple hexose sugars including glucose do not stimulate its growth, even on defined medium. However, M. xanthus is sensitive to the glucose analogue, 2-deoxyglucose (2dGIc). This antibiotic is toxic, because it is phosphorylated by hexokinase, the first enzyme in the primary glycolytic pathway for sugar catabolism present in most organisms that utilize glucose. The isolation and characterization of spontaneous mutants of M. xanthus resistant to 2dGIc shows that M. xanthus makes a hexokinase and must transport simple sugars. This hexokinase is unusual, because it is inhibited by the two essential hexosamine sugars glucosamine and N-acetylglucosamine. Another bacterial hexokinase, made by E. coil, is also inhibited by the same sugars. The three main goals of this research are: (1) to understand why these enzymes are sensitive to 2dGIc and inhibited by hexosamines; (2), to identify the precise target of 2dGIc-6-phosphate responsible for the toxicity of 2dGIc in most organisms; and (3) to understand the regulatory mechanism in this organism that governs the preference for amino acids over glucose. To accomplish the first task, the kinetics of substrate binding and utilization of these sugars by hexokinase will be examined using kinetic and mechanistic approaches. The hexokinase gene from M. xanthus will be cloned and sequenced, and both the M. xanthus and E. coil enzymes will be purified. Second, the recent discovery that the production of intracellular 2dGIc-6-phosphate results in the depletion of ATP in M. xanthus, as it does in mammalian cells, has led to the search for the critical enzyme involved in central carbon metabolism specifically inhibited by low concentrations of 2dGIc-6-phosphate, again using traditional biochemical tactics. Third, glucose catabolism in M. virescens, which oxidizes glucose readily, and in M. xanthus, which may not, will be compared.Many living organisms, like humans, burn sugars for energy, but prefer a rich protein diet. The soil bacterium, Myxococcus xanthus, chooses a protein-rich diet ("steak") over a carbohydrate-rich diet ("potatoes") to an extreme, because sugar doesn't stimulate its growth. The goal of this research is to understand why some individual organisms have this kind of dietary preference. Biochemical studies show that the steak-loving bacterium M. xanthus can still taste sugars, because it is fooled by the sweet poison of 2-deoxyglucose, which tastes like sweet glucose-rich Gatorade TM, but has the opposite effect. This poison, 2-deoxyglucose, has been known to be toxic to most living things for 50 years. No one really understands why, except for the finding that it drains ATP (energy) levels in many living cells, including M. xanthus. M. xanthus will be used as a model organism to figure out why this poison is a poison, and, at the same time, gain fundamental insights at a basic chemical level about how organisms choose what to eat.