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Although genome-editing enzymes such as TALEN and CRISPR/Cas9 are being widely used, they have an essential limitation in that their relatively high molecular weight makes them difficult to be delivered to cells. To develop a novel genome-editing enzyme with a smaller molecular weight, we focused on the engrailed homeodomain (EHD). We designed and constructed proteins composed of two EHDs connected by a linker to increase sequence specificity. In bacterial one-hybrid assays and EMSA analyses, the created proteins exhibited good affinity for DNA sequences consisting of two tandemly aligned EHD target sequences. However, they also bound to individual EHD targets. To avoid binding to single target sites, we introduced amino acid mutations to reduce the protein-DNA affinity of each EHD monomer and successfully created a small protein with high specificity for tandem EHD target sequences. This article is protected by copyright. All rights reserved.
This article was published in the following journal.
Name: Protein science : a publication of the Protein Society
Selective autophagy enables degradation of specific cargo such as protein aggregates or organelles and thus plays an essential role in the regulation of cellular homeostasis. Cargo specificity is achi...
Aptamers are nucleic acid molecules that bind to a target molecule with high affinity and specificity, which are generated by a process known as systematic evolution of ligands by exponential enrichme...
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A family of RNA-BINDING PROTEINS that contain an RNA RECOGNITION MOTIF and two ribonucleoprotein (RNP) domains which bind RNA, in addition to other domains that allow for high affinity binding, sequence specificity, and protein interactions. Examples of RNA recognition motif proteins include HETEROGENEOUS NUCLEAR RIBONUCLEARPROTEINS (hnRNP) and EMBRYONIC LETHAL ABNORMAL-VISION (ELAV) proteins.
The property of antibodies which enables them to react with some ANTIGENIC DETERMINANTS and not with others. Specificity is dependent on chemical composition, physical forces, and molecular structure at the binding site.
An approximately 80 amino acid RNA binding motif that consists of four anti-parallel surface beta sheets and two alpha helices arranged in a beta-alpha-beta-beta-alpha-beta configuration. One of the surface beta sheets interacts with two or three specific RNA bases. Interactions between additional sequences and the RNA, as well as within the RNA recognition motif increase the affinity and specificity of the protein-RNA interaction.
The interaction of two or more substrates or ligands with the same binding site. The displacement of one by the other is used in quantitative and selective affinity measurements.
Homeostatic control of the immune system by secretion of different cytokines by the Th1 and Th2 cells. The concentration dependent binding of the various cytokines to specific receptors determines the balance (or imbalance leading to disease).
Bioinformatics is the application of computer software and hardware to the management of biological data to create useful information. Computers are used to gather, store, analyze and integrate biological and genetic information which can then be applied...
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Immunoassay - ELISA
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