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

过去的几十年见证了分子生物学领域的诞生和成熟。最初，突变体 基因是基因组研究的焦点。现在，整个基因组都被例行测序，而 通过注解算法自动识别驻留基因。或者，蛋白质组学方法 制备全细胞提取液中的蛋白水解肽，用于质谱学分析。这其中的每一个 方法对大基因有很强的偏见：大基因是经常突变的目标，长期开放- 基因组序列中的阅读框很容易辨认，大的蛋白质产生许多多肽 用于质谱学鉴定。这种无意的偏见也在我们的 对分子生物学的理解。 最近在真核生物和原核生物中的工作已经发现了大量的小基因或它们的 编码的蛋白质。小蛋白质的数量(被认为是50aa或更少)可以与传统的 大的蛋白质，但只有少数被认为是一种功能。这项提议的目标是推动 这一新兴领域通过促进小蛋白的发现和功能表征而向前发展。我们 将在分枝杆菌中开发和应用我们的小蛋白方法。分枝杆菌提供了许多 小蛋白研究的优势。最重要的是，它们以标准的方式表达&gt；1000小蛋白 条件。 本增刊的目的是为了支持一位杰出的本科生劳伦·福克斯，她渴望 成为一名研究科学家，具有直接动手的实验室研究经验。劳伦将专注于 剖析一组独特的半胱氨酸蛋白介导的调控机制 动态平衡。她将学习新的技术和分析技能，并加深对 成为一名研究科学家的必备条件。参与沃兹沃斯夏季研究计划 本科生将进一步让劳伦接触到潜在的科学职业、未来的导师和新朋友 和同事们。

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.