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Title: Altering the orientation of a fused protein to the RNA-binding ribosomal protein L7Ae and its derivatives through circular permutation

Abstract

RNA-protein complexes (RNPs) are useful for constructing functional nano-objects because a variety of functional proteins can be displayed on a designed RNA scaffold. Here, we report circular permutations of an RNA-binding protein L7Ae based on the three-dimensional structure information to alter the orientation of the displayed proteins on the RNA scaffold. An electrophoretic mobility shift assay and atomic force microscopy (AFM) analysis revealed that most of the designed circular permutants formed an RNP nano-object. Moreover, the alteration of the enhanced green fluorescent protein (EGFP) orientation was confirmed with AFM by employing EGFP on the L7Ae permutant on the RNA. The results demonstrate that targeted fine-tuning of the stereo-specific fixation of a protein on a protein-binding RNA is feasible by using the circular permutation technique.

Authors:
;  [1];  [2];  [1]
  1. Graduate School of Biostudies, Kyoto University, Oiwake-cho, Kitashirakawa, Sakyo-ku, Kyoto, 606-8502 (Japan)
  2. Department of Life and Environmental Sciences, Faculty of Engineering, Chiba Institute of Technology, 2-17-1 Tsudanuma, Narashino, Chiba, 275-0016 (Japan)
Publication Date:
OSTI Identifier:
22592766
Resource Type:
Journal Article
Resource Relation:
Journal Name: Biochemical and Biophysical Research Communications; Journal Volume: 466; Journal Issue: 3; Other Information: Copyright (c) 2015 Elsevier Science B.V., Amsterdam, The Netherlands, All rights reserved.; Country of input: International Atomic Energy Agency (IAEA)
Country of Publication:
United States
Language:
English
Subject:
60 APPLIED LIFE SCIENCES; ATOMIC FORCE MICROSCOPY; FLUORESCENCE; INFORMATION; PROTEINS; RNA

Citation Formats

Ohuchi, Shoji J., Sagawa, Fumihiko, Sakamoto, Taiichi, and Inoue, Tan, E-mail: tan@kuchem.kyoto-u.ac.jp. Altering the orientation of a fused protein to the RNA-binding ribosomal protein L7Ae and its derivatives through circular permutation. United States: N. p., 2015. Web. doi:10.1016/J.BBRC.2015.09.035.
Ohuchi, Shoji J., Sagawa, Fumihiko, Sakamoto, Taiichi, & Inoue, Tan, E-mail: tan@kuchem.kyoto-u.ac.jp. Altering the orientation of a fused protein to the RNA-binding ribosomal protein L7Ae and its derivatives through circular permutation. United States. doi:10.1016/J.BBRC.2015.09.035.
Ohuchi, Shoji J., Sagawa, Fumihiko, Sakamoto, Taiichi, and Inoue, Tan, E-mail: tan@kuchem.kyoto-u.ac.jp. Fri . "Altering the orientation of a fused protein to the RNA-binding ribosomal protein L7Ae and its derivatives through circular permutation". United States. doi:10.1016/J.BBRC.2015.09.035.
@article{osti_22592766,
title = {Altering the orientation of a fused protein to the RNA-binding ribosomal protein L7Ae and its derivatives through circular permutation},
author = {Ohuchi, Shoji J. and Sagawa, Fumihiko and Sakamoto, Taiichi and Inoue, Tan, E-mail: tan@kuchem.kyoto-u.ac.jp},
abstractNote = {RNA-protein complexes (RNPs) are useful for constructing functional nano-objects because a variety of functional proteins can be displayed on a designed RNA scaffold. Here, we report circular permutations of an RNA-binding protein L7Ae based on the three-dimensional structure information to alter the orientation of the displayed proteins on the RNA scaffold. An electrophoretic mobility shift assay and atomic force microscopy (AFM) analysis revealed that most of the designed circular permutants formed an RNP nano-object. Moreover, the alteration of the enhanced green fluorescent protein (EGFP) orientation was confirmed with AFM by employing EGFP on the L7Ae permutant on the RNA. The results demonstrate that targeted fine-tuning of the stereo-specific fixation of a protein on a protein-binding RNA is feasible by using the circular permutation technique.},
doi = {10.1016/J.BBRC.2015.09.035},
journal = {Biochemical and Biophysical Research Communications},
number = 3,
volume = 466,
place = {United States},
year = {Fri Oct 23 00:00:00 EDT 2015},
month = {Fri Oct 23 00:00:00 EDT 2015}
}