Department of Chemistry and Chemical Biology, Harvard University, 12 Oxford Street, Cambridge, MA 02138, USA.
Department of Chemistry, Merkert Chemistry Center, Boston College, Chestnut Hill, Massachusetts 02467, USA.
J Am Chem Soc. 2013 Nov 13;135(45):16750-3. doi: 10.1021/ja406606j. Epub 2013 Oct 25.
The application of ribosome profiling and mass spectrometry technologies has recently revealed that the human proteome is larger than previously appreciated. Short open reading frames (sORFs), which are difficult to identify using traditional gene-finding algorithms, constitute a significant fraction of unknown protein-coding genes. Thus, experimental approaches to identify sORFs provide invaluable insight into the protein-coding potential of genomes. Here, we report an affinity-based approach to enrich and identify cysteine-containing human sORF-encoded polypeptides (ccSEPs) from cells. This approach revealed 16 novel ccSEPs, each derived from an uncharacterized sORF, demonstrating its potential for discovering new genes. We validated expression of a SEP from its endogenous RNA, and demonstrated the specificity of our labeling approach using synthetic SEP. The discovery of additional human SEPs and their conservation indicate the potential importance of these molecules in biology.
核糖体图谱和质谱技术的应用最近揭示，人类蛋白质组比以前认为的要大。使用传统基因发现算法难以识别的短开放阅读框（sORF）构成了未知蛋白质编码基因的重要部分。因此，鉴定 sORF 的实验方法为研究基因组的蛋白质编码潜力提供了宝贵的见解。在这里，我们报告了一种基于亲和性的方法，从细胞中富集和鉴定含半胱氨酸的人 sORF 编码多肽（ccSEP）。该方法揭示了 16 个新的 ccSEP，每个都来自一个未被表征的 sORF，证明了它发现新基因的潜力。我们从其内源性 RNA 验证了 SEP 的表达，并使用合成 SEP 证明了我们标记方法的特异性。额外的人类 SEP 的发现及其保守性表明这些分子在生物学中具有潜在的重要性。
