The presence of a bacterial metabolic pathway in eukaryotic fungi

描述（由申请人提供）：已在真核真菌中鉴定出由乙酸激酶（ACK）和木酮糖5-磷酸/果糖6-磷酸磷酸转酮酶（XFP）组成的细菌代谢途径。在细菌中，ACK主要与其伴侣酶磷酸转乙酰酶（PTA）一起用于乙酸和乙酰辅酶A的相互转化。然而，PTA在真菌中不存在。相反，ACK与XFP形成了一个假定的途径，XFP和ACK一样，也被认为几乎严格属于细菌。已经提出ACK-XFP形成乳酸菌异型发酵生长中的修饰的戊糖磷酸转酮酶途径的一部分，并且在能量产生中起关键作用，但是该途径在真菌中的作用尚未被研究。 事实上，所有关于ACK酶学的知识都来自于对E.大肠杆菌ACK和唯一的古细菌ACK，来自甲烷八叠球菌。最初的生化和动力学特性的几个真核生物的ACK揭示了有趣的和独特的性能，这表明这种酶的家庭是显着更多样化比以前认为的。这些生化差异表明，真核生物的ACK可能有不同的生理功能比他们的原核生物。对XFP的生物化学和酶学知之甚少。令人惊讶的是，许多真菌有两个基因编码这种酶，提高了可能性，编码的酶之间的差异是生物化学和生理学的重要性。 本研究的目的是了解假定的ACK-XFP途径在真菌生理和代谢中的作用，使用遗传上易处理的担子菌新生隐球菌作为模型系统。该提议的具体目的是双重的：（1）研究ACK-XFP途径在真菌代谢和生理学中的作用，以及（2）生物化学和动力学表征真菌ACK和XFP酶。 这将是首次对真菌中ACK-XFP途径的生理作用进行研究，也是首次对主要不与PTA合作的ACK进行研究。本提案中描述的实验有望为这一重要代谢酶的酶学和进化提供独特的见解，并有助于更深入地了解磷酸转移酶的ASKHA（乙酸和糖激酶/Hsc 70/肌动蛋白）超家族。 公共卫生相关性：最近在许多医学和经济相关的真菌中发现了由乙酸激酶（ACK）和木酮糖5-磷酸/果糖6-磷酸转酮酶（XFP）组成的细菌能量产生途径。如果ACK和XFP在真菌中被证明是必不可少的，那么它们在人类、动物和植物中的缺失可能会使这些酶成为未来开发抗真菌剂的潜在靶点。然而，在考虑这种努力之前，彻底了解真菌中ACK-XFP途径的生物化学和生理作用是必要的。

DESCRIPTION (provided by applicant): A bacterial metabolic pathway composed of acetate kinase (ACK) and xylulose 5- phosphate/fructose 6-phosphate phosphoketolase (XFP) has been identified in eukaryotic fungi. In bacteria, ACK primarily functions with its partner enzyme phosphotransacetylase (PTA) for the interconversion of acetate and acetyl-CoA. However, PTA is absent in fungi. Instead, ACK forms a putative pathway with XFP, which, like ACK, was also thought to belong almost strictly to the bacteria. ACK-XFP has been proposed to form part of a modified pentose phosphoketolase pathway in the heterofermentative growth of lactic acid bacteria and play a key role in energy generation, but the role of this pathway in fungi has not yet been investigated. Virtually everything known about the enzymology of ACK is from characterization of the E. coli ACK and the only archaeal ACK, that from Methanosarcina. Initial biochemical and kinetic characterization of several eukaryotic ACKs has revealed interesting and unique properties, suggesting that this enzyme family is significantly more diverse than previously thought. These biochemical differences indicate that eukaryotic ACKs may have different physiological functions than their prokaryotic counterparts. Very little is known about the biochemistry and enzymology of XFP. Surprisingly, a number of fungi have two genes encoding this enzyme, raising the possibility that the differences between the encoded enzymes is of biochemical and physiological importance. The objective of this study is to understand the role of the putative ACK-XFP pathway in fungal physiology and metabolism using the genetically tractable basidiomycete Cryptococcus neoformans as the model system. The specific aims of this proposal are two-fold: (1) to investigate the role of the ACK-XFP pathway in fungal metabolism and physiology, and (2) to biochemically and kinetically characterize fungal ACK and XFP enzymes. This will be the first investigation into the physiological role of the ACK-XFP pathway in fungi as well as the first into ACKs that do not primarily partner with PTA. The experiments described in this proposal are expected to provide unique insights into the enzymology and evolution of this important metabolic enzyme as well as contribute to a deeper understanding of the ASKHA (acetate and sugar kinase/Hsc70/actin) superfamily of phosphotransferases. PUBLIC HEALTH RELEVANCE: A bacterial energy generating pathway consisting of the enzymes acetate kinase (ACK) and xylulose 5-phosphate/fructose 6-phosphate phosphoketolase (XFP) has recently been found in a number of medically and economically relevant fungi. If ACK and XFP prove to be essential in fungi, their absence in humans, animals, and plants may allow these enzymes to be investigated as potential targets for development of antifungal agents in the future. However, a thorough understanding of the biochemistry and physiological role of the ACK-XFP pathway in fungi, the objectives of this proposal, is necessary before such endeavors can be considered.