文章基本信息

标题：Mutant libraries reveal negative design shielding proteins from supramolecular self-assembly and relocalization in cells
本地全文：下载
作者：Hector Garcia Seisdedos ; Tal Levin ; Gal Shapira 等
期刊名称：Proceedings of the National Academy of Sciences
印刷版ISSN：0027-8424
电子版ISSN：1091-6490
出版年度：2022
卷号：119
期号：5
DOI：10.1073/pnas.2101117119
语种：English
出版社：The National Academy of Sciences of the United States of America
摘要：Significance Genetic mutations fuel organismal evolution but can also cause disease. As proteins are the cell’s workhorses, the ways in which mutations can disrupt their structure, stability, function, and interactions have been studied extensively. However, proteins evolve and function in a cellular context, and our ability to relate changes in protein sequence to cell-level phenotypes remains limited. In particular, the molecular mechanism underlying most disease-associated mutations is unknown. Here, we show that mutations changing a protein’s surface chemistry can dramatically impact its supramolecular self-assembly and localization in the cell. These results highlight the complex nature of genotype–phenotype relationships with a simple system. Understanding the molecular consequences of mutations in proteins is essential to map genotypes to phenotypes and interpret the increasing wealth of genomic data. While mutations are known to disrupt protein structure and function, their potential to create new structures and localization phenotypes has not yet been mapped to a sequence space. To map this relationship, we employed two homo-oligomeric protein complexes in which the internal symmetry exacerbates the impact of mutations. We mutagenized three surface residues of each complex and monitored the mutations’ effect on localization and assembly phenotypes in yeast cells. While surface mutations are classically viewed as benign, our analysis of several hundred mutants revealed they often trigger three main phenotypes in these proteins: nuclear localization, the formation of puncta, and fibers. Strikingly, more than 50% of random mutants induced one of these phenotypes in both complexes. Analyzing the mutant’s sequences showed that surface stickiness and net charge are two key physicochemical properties associated with these changes. In one complex, more than 60% of mutants self-assembled into fibers. Such a high frequency is explained by negative design: charged residues shield the complex from self-interacting with copies of itself, and the sole removal of the charges induces its supramolecular self-assembly. A subsequent analysis of several other complexes targeted with alanine mutations suggested that such negative design is common. These results highlight that minimal perturbations in protein surfaces’ physicochemical properties can frequently drive assembly and localization changes in a cellular context.
关键词：enprotein evolutionprotein interactionsgenotype–phenotype map