Biosynthesis of hypermodified guanosines

描述（由申请人提供）：项目概述。转移RNA （tRNA）的转录后加工涉及许多功能不同的事件，这些事件对tRNA成熟至关重要。核苷修饰现象可能是这些事件中最引人注目的，它会导致典型核苷的大量结构变化。在tRNA中发现的两个最显著的修饰核苷是7-去氮杂鸟苷衍生物queuosine和archaeosine，它们分别在翻译和RNA稳定中发挥作用。虽然进化相关，但这些核苷在不同的结构域内分离；细菌和真核生物中普遍存在排队苷，而古细菌中只存在古细菌。一般来说，7-去氮杂嘌呤结构在生物学中广泛存在，它存在于各种天然产物中，如链霉菌的抗肿瘤抗生素toyocamycin， sangivamycin和结核菌素。对这些去氮嘌呤的生物合成途径了解甚少，这一事实阻碍了功能研究。数百个测序基因组的可用性现在允许使用比较基因组学方法识别基因和途径。利用这种方法发现了5种新的酶，这些酶可用于重新合成队列苷和古苷，以及其他7-去氮杂嘌呤代谢物。值得注意的是，这一途径仅限于原核生物，其中一些新发现的酶似乎催化了生物学上前所未有的化学反应。本项目的长期目标是阐明7-去氮嘌呤在原核生物中的生物合成和代谢。本研究的具体目的是：1)阐明这些新酶在导致共同前体7-氰-7-去氮鸟嘌呤形成的队列苷和古苷生物合成途径的早期步骤中的作用；2)启动对这些修饰核苷的更广泛代谢的研究；3)研究其中一种酶的机制和结构，一种新的腈氧化还原酶。这一建议带来了生物信息学，遗传学，生化和化学方法的集合来阐明7-去氮杂鸟嘌呤修饰核苷的生物合成问题。这一新途径及其组成酶的研究将为阐明7-去氮杂嘌呤代谢及其调控的生物学提供前所未有的途径。的相关性。7-去氮嘌呤的途径是微生物所特有的，许多构成酶是潜在的新的抗菌靶点。此外，其中一种酶是一种新型的腈氧化还原酶，可能在工业生物催化中有应用。

DESCRIPTION (provided by applicant): Project Summary. The post-transcriptional processing of transfer RNA (tRNA) involves a number of functionally distinct events essential for tRNA maturation. The phenomenon of nucleoside modification is perhaps the most remarkable of these events, and results in a wealth of structural changes to the canonical nucleosides. Two of the most remarkable modified nucleosides found in tRNA are the 7-deazaguanosine derivatives queuosine and archaeosine, which have putative roles in translation and RNA stabilization, respectively. While evolutionarily related, these nucleosides are segregated within separate Domains; queuosine is ubiquitous among Bacteria and Eukarya, while archaeosine is only present in the Archaea. The 7-deazapurine structure in general is widespread in biology, where it is found in a variety of natural products such as the antitumor antibiotics toyocamycin, sangivamycin, and tubercidin from Streptomyces. The biosynthetic pathways to these deazapurines are poorly understood, a fact that has stymied functional studies. The availability of hundreds of sequenced genomes now allows the identification of genes and pathways using a comparative genomics approach. This approach was used to discover five new enzymes in the de novo biosynthesis of queuosine and archaeosine, and potentially of other 7-deazapurine metabolites. Notably, this pathway is limited to prokaryotes, and some of these newly discovered enzymes appear to catalyze chemistry unprecedented in biology. The long-term objectives of this project are to elucidate the biosynthesis and metabolism of 7-deazapurines in prokaryotes. The specific aims of this proposal are 1) to elucidate the role of these new enzymes in the early steps in the queuosine and archaeosine biosynthetic pathways leading to the formation of the common precursor 7-cyano-7-deazaguanine, 2) to initiate studies into the broader metabolism of these modified nucleosides, and 3) to investigate the mechanism and structure of one of these enzymes, a novel nitrile oxidoreductase. This proposal brings an ensemble of bioinformatic, genetic, biochemical, and chemical approaches to the problem of elucidating the biosynthesis of 7-deazaguanine modified nucleosides. The study of this new pathway and the constituent enzymes will provide unprecedented access to elucidating the biology of 7-deazapurine metabolism and its manipulation. Relevance. The pathway to 7-deazapurines is unique to microorganisms, and many of the constitute enzymes are potentially new antibacterial targets. Furthermore, one of the enzymes is a novel nitrile oxidoreductase that may have applications in industrial biocatalysis.