Identification of Microprotein-Protein Interactions via APEX Tagging

被引:46
作者
Chu, Qian [1 ]
Rathore, Annie [1 ,2 ]
Diedrich, Jolene K. [1 ,3 ]
Donaldson, Cynthia J. [1 ]
Yates, John R., III [3 ]
Saghatelian, Alan [1 ]
机构
[1] Salk Inst Biol Studies, Clayton Fdn Labs Peptide Biol, 10010 North Torrey Pines Rd, La Jolla, CA 92037 USA
[2] Univ Calif San Diego, Div Biol Sci, 9500 Gilman Dr, La Jolla, CA 92093 USA
[3] Scripps Res Inst, Dept Physiol Chem, 10550 North Torrey Pines Rd, La Jolla, CA 92037 USA
关键词
OPEN READING FRAMES; LIVING CELLS; MESSENGER-RNA; PEPTIDES; NUCLEOPHOSMIN; DISCOVERY; SIGNAL; POLYPEPTIDES; DROSOPHILA; NETWORK;
D O I
10.1021/acs.biochem.7b00265
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Microproteins are peptides and small proteins encoded by small open reading frames (smORFs). Newer technologies have led to the recent discovery of hundreds to thousands of new microproteins. The biological functions of a few inicropioteins have been elucidated, and these micro-proteins have fundamental roles in biology ranging from limb development to muscle function, highlighting the Value of characterizing these molecules. The identification of microprotein protein interactions,(MPIs) has proven to be a successful approach to the functional characterization of these genes; however, traditional immunoprecipitation methods result in the, enrichment of nonspecific. interactions for, microproteins. Here, we test and apply an in situ proximity tagging method that relies on an engineered ascorbate peroxidase 2 (APEX) to elucidate MPls. The results,demonstrate that APEX tagging is superior. to traditional immunoprecipitation methods for microproteins. Furthermore,the application of APEX tagging to an uncharacterized microprotein called Cllorf98 revealed that this microprotein interacts with nucleolar proteins nucleophosmin and nucleolin, demonstrating the ability,of this approach to identify novel. hypothesis-generating MPIs.
引用
收藏
页码:3299 / 3306
页数:8
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