A variant of green fluorescent protein exclusively deposited to active intracellular inclusion bodies

Microbial Cell Factories

Research

Hyojin Moon¹  Sebyung Kang¹  Seong Soo A An²  Sun-Gu Lee³  Govindan Raghunathan³  Ganapathiraman Munussami³  Yong-Sung Kim⁴  Hyun-jong Paik⁵ 

[1]Department of Biological Sciences, School of Life Sciences, Ulsan National Institute of Science and Technology (UNIST), Ulsan, South Korea

[2]Department of Bionanotechnology, Gachon University, Seongnam, South Korea

[3]Department of Chemical Engineering, Pusan National University, Busan, South Korea

[4]Department of Molecular Science and Technology, Ajou University, Suwon, South Korea

[5]Department of Polymer Science and Engineering, Pusan National University, Busan, South Korea

关键词: Green fluorescent protein;    Active inclusion bodies;    Protein aggregation;    Fluorescent particle;    Protein folding;   

DOI  :  10.1186/1475-2859-13-68

 received in 2014-01-13, accepted in 2014-05-11,  发布年份 2014

来源: Springer

【 摘 要 】

BackgroundInclusion bodies (IBs) were generally considered to be inactive protein deposits and did not hold any attractive values in biotechnological applications. Recently, some IBs of recombinant proteins were confirmed to show their functional properties such as enzyme activities, fluorescence, etc. Such biologically active IBs are not commonly formed, but they have great potentials in the fields of biocatalysis, material science and nanotechnology.ResultsIn this study, we characterized the IBs of DL4, a deletion variant of green fluorescent protein which forms active intracellular aggregates. The DL4 proteins expressed in Escherichia coli were exclusively deposited to IBs, and the IBs were estimated to be mostly composed of active proteins. The spectral properties and quantum yield of the DL4 variant in the active IBs were almost same with those of its native protein. Refolding and stability studies revealed that the deletion mutation in DL4 didn’t affect the folding efficiency of the protein, but destabilized its structure. Analyses specific for amyloid-like structures informed that the inner architecture of DL4 IBs might be amorphous rather than well-organized. The diameter of fluorescent DL4 IBs could be decreased up to 100–200 nm by reducing the expression time of the protein in vivo.ConclusionsTo our knowledge, DL4 is the first GFP variant that folds correctly but aggregates exclusively in vivo without any self-aggregating/assembling tags. The fluorescent DL4 IBs have potentials to be used as fluorescent biomaterials. This study also suggests that biologically active IBs can be achieved through engineering a target protein itself.

【 授权许可】

Unknown   
© Raghunathan et al.; licensee BioMed Central Ltd. 2014. This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly credited. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.