Systematic Discovery and Analysis of Small Proteins and Small ORFs in Mycobacteria

过去几十年见证了分子生物学领域的诞生和成熟。最初，突变 基因是基因组探索的焦点。现在，整个基因组都被例行测序， 通过注释算法自动识别常驻基因。另外，蛋白质组学方法 制备用于质谱分析的全细胞提取物的蛋白水解肽。这一切成功都 方法对大基因有强烈的偏见：大基因是突变的常见目标，长期开放， 阅读框在基因组序列中很容易识别，大的蛋白质产生许多肽 用于质谱鉴定。这种无意的偏见也造成了我们的巨大差距， 了解分子生物学。 最近在真核生物和原核生物中的研究发现了大量的小基因或其 编码蛋白质。小分子蛋白质（被认为是50个氨基酸或更少）的数量与传统的 大的蛋白质，但只有少数已被归因于一个功能。该提案的目的是推动 这一新兴领域的发展，促进了小蛋白的发现和功能表征。我们 将在分枝杆菌中开发和应用我们的小蛋白方法。分枝杆菌提供许多 小蛋白研究的优势。最重要的是，它们在标准蛋白质中表达>1000种小蛋白质。 条件 这个补充的目标是支持一个优秀的本科生，劳伦福克斯，谁渴望 成为一名研究科学家，拥有直接的实验室研究经验。劳伦会专注于 剖析半胱氨酸中一组独特的s蛋白介导的调节机制 体内平衡她将学习新的技术和分析技能，并更好地了解 成为一名研究科学家需要什么参与沃兹沃斯夏季研究计划， 本科生将进一步暴露劳伦在科学，未来的导师和新朋友的潜在职业 和同事。

The last few decades have seen the birth and maturation of the field of Molecular Biology. Initially, mutant genes were focal points of genome exploration. Now, entire genomes are routinely sequenced, and the resident genes are automatically identified by annotation algorithms. Alternatively, proteomic approaches prepare proteolytic peptides of whole-cell extracts for analysis by mass spectrometry. Each of these approaches are strongly biased for large genes: large genes are frequent targets for mutation, long-open- reading frames are easily discerned in genomic sequence, and large proteins generate many peptides for mass spectrometry identification. This unintended bias has also created a large gap in our understanding of molecular biology. Recent work in eukaryotes and prokaryotes alike have uncovered multitudes of small genes or their encoded proteins. The numbers of small proteins (considered as 50 aa or less) rival that of traditionally large proteins, yet only a handful have been ascribed a function. The goal of this proposal is to propel this nascent field forward by facilitating both small protein discovery and functional characterization. We will develop and apply our small protein approaches in mycobacteria. Mycobacteria offer many advantages for small protein study. Foremost is that they express >1000 small proteins in standard conditions. The goal of this supplement is to support an outstanding undergraduate student, Lauren Fox, who aspires to be a research scientist, with direct hands-on laboratory research experience. Lauren will focus on dissecting the mechanism of regulation mediated by a unique set of sproteins involved in cysteine homeostasis. She will learn new technical and analytical skills, and develop a greater understanding of what it takes to be a research scientist. Involvement in the Wadsworth summer research program for undergraduates will further expose Lauren to potential careers in science, future mentors and new friends and colleagues.