Pyrrolysine, a novel genetically encoded amino acid

描述（由申请人提供）：吡咯赖氨酸是一种新型氨基酸，最近发现由微生物甲胺代谢基因中的UAG密码子编码。这是继18年前发现由微生物中的UGA密码子编码的硒代半胱氨酸之后，基因编码的非规范氨基酸的第二个例子。硒代半胱氨酸目前已知分布广泛，是人体中硒的主要形式。根据这一先例，吡咯赖氨酸具有更广泛分布和代谢重要性的潜力。无论如何，吡咯赖氨酸将作为一个罕见的例子，说明生物体如何调节其遗传密码以扩大代谢能力，这一成就的复制有望为人工定制具有生物医学潜力的新型蛋白质。我们的长期目标是了解吡咯赖氨酸的功能，生物起源和遗传编码。假设UAG密码子的翻译需要专门的UAG解码tRNA和专用的同源氨酰-tRNA合成酶。这些基因产物和其他被认为参与UAG解码的吡咯赖氨酸的作用将使用生物化学和遗传学方法进行研究。吡咯赖氨酸的最终结构尚不清楚，通过分析蛋白质、胞质溶胶和tRNA上的UAG编码残基，将深入了解吡咯赖氨酸的结构和遗传编码。已经开发了一种体内系统，该系统将用于研究UAG在作为吡咯赖氨酸的UAG翻译期间如何作为有义密码子而不是终止密码子起作用。一个类似的系统将用于定点诱变，以测试吡咯赖氨酸在酶催化中的假设功能。最后，转座子诱变将用于鉴定在作为吡咯赖氨酸的UAG翻译期间具有预期或非预期功能的未知基因。

DESCRIPTION (provided by applicant): Pyrrolysine, a novel amino acid, was recently found to be encoded by UAG codons in a microbial gene for methylamine metabolism. This is the second example of a genetically encoded non-canonical amino acid since selenocysteine, which 18 years ago was found to be encoded by UGA codons in a microbe. Selenocysteine is now known to be widely distributed and the primary form of selenium in humans. Following this precedent, pyrrolysine holds potential of being more widely distributed and of import in metabolism. Regardless, pyrrolysine will serve as a rare example of how organisms can modulate their genetic code to expand metabolic capabilities, an achievement whose replication holds promise for artificially tailoring novel proteins with biomedical potential. Our long-range goals are to understand the function, biogenesis, and genetic encoding of pyrrolysine. Translation of UAG codons is hypothesized to require a specialized UAG decoding tRNA and a dedicated cognate aminoacyl-tRNA synthetase. The roles of these gene products and others thought to be involved in UAG decoding as pyrrolysine will be investigated using biochemical and genetic approaches. The final structure of pyrrolysine is as yet unknown, and insight into pyrrolysine structure and genetic encoding will be gained by analyzing the UAG encoded residue in protein, the cytosol, and on tRNA. An in vivo system has been developed that will be used to study how UAG functions as a sense rather than a stop codon during UAG translation as pyrrolysine. A similar system will be used for site directed mutagenesis in order to test the hypothesized function of pyrrolysine in enzyme catalysis. Finally, transposon mutagenesis will be used to identify unknown genes with anticipated or unanticipated functions during UAG translation as pyrrolysine.