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Production of bioactive chicken (Gallus gallus) follistatin-type proteins in E. coli

Follistatin (FST) is a cysteine-rich autocrine glycoprotein and plays an important role in mammalian prenatal and postnatal development. FST binds to and inhibit myostatin (MSTN), a potent negative regulator of skeletal muscle growth, and FST abundance enhances muscle growth in animals via inhibitio...

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Autores principales: Lee, Sang Beum, Park, Sung Kwon, Kim, Yong Soo
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Springer Berlin Heidelberg 2015
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4547976/
https://www.ncbi.nlm.nih.gov/pubmed/26302688
http://dx.doi.org/10.1186/s13568-015-0142-3
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author Lee, Sang Beum
Park, Sung Kwon
Kim, Yong Soo
author_facet Lee, Sang Beum
Park, Sung Kwon
Kim, Yong Soo
author_sort Lee, Sang Beum
collection PubMed
description Follistatin (FST) is a cysteine-rich autocrine glycoprotein and plays an important role in mammalian prenatal and postnatal development. FST binds to and inhibit myostatin (MSTN), a potent negative regulator of skeletal muscle growth, and FST abundance enhances muscle growth in animals via inhibition of MSTN activity. The objective of this study was to produce biologically active, four chicken FST-type proteins in an Escherichia coli expression system. Gibson assembly cloning method was used to insert the DNA fragments of four FST-type proteins, designated as FST288, NDFSD1/2, NDFSD1, and NDFSD1/1, into pMALc5x vector downstream of the maltose-binding protein (MBP) gene, and the plasmids containing the inserts were eventually transformed into Shuffle E. coli strain for protein expression. We observed a soluble expression of the four MBP-fused FST-type proteins, and the proteins could be easily purified by the combination of amylose and heparin resin affinity chromatography. MBP-fused FST-type proteins demonstrated their affinity to anti-FST antibody. In an in vitro reporter gene assay to examine their potencies and selectivities to different ligands (MSTN, GDF11, and activin A), the four FST-type proteins (MBP-FST288, MBP-NDFSD1/2, MBP-NDFSD1, and MBP-NDFSD1/1) showed different potency and selectivity against the three ligands from each other. Ligand selectivity of each FST-type proteins was similar to its counterpart FST-type protein of eukaryotic origin. In conclusion, we could produce four FST-type proteins having different ligand selectivity in E. coli, and the results imply that economic production of a large amount of FST-type proteins with different ligand selectivity is possible to examine their potential use in meat-producing animals.
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spelling pubmed-45479762015-08-28 Production of bioactive chicken (Gallus gallus) follistatin-type proteins in E. coli Lee, Sang Beum Park, Sung Kwon Kim, Yong Soo AMB Express Original Article Follistatin (FST) is a cysteine-rich autocrine glycoprotein and plays an important role in mammalian prenatal and postnatal development. FST binds to and inhibit myostatin (MSTN), a potent negative regulator of skeletal muscle growth, and FST abundance enhances muscle growth in animals via inhibition of MSTN activity. The objective of this study was to produce biologically active, four chicken FST-type proteins in an Escherichia coli expression system. Gibson assembly cloning method was used to insert the DNA fragments of four FST-type proteins, designated as FST288, NDFSD1/2, NDFSD1, and NDFSD1/1, into pMALc5x vector downstream of the maltose-binding protein (MBP) gene, and the plasmids containing the inserts were eventually transformed into Shuffle E. coli strain for protein expression. We observed a soluble expression of the four MBP-fused FST-type proteins, and the proteins could be easily purified by the combination of amylose and heparin resin affinity chromatography. MBP-fused FST-type proteins demonstrated their affinity to anti-FST antibody. In an in vitro reporter gene assay to examine their potencies and selectivities to different ligands (MSTN, GDF11, and activin A), the four FST-type proteins (MBP-FST288, MBP-NDFSD1/2, MBP-NDFSD1, and MBP-NDFSD1/1) showed different potency and selectivity against the three ligands from each other. Ligand selectivity of each FST-type proteins was similar to its counterpart FST-type protein of eukaryotic origin. In conclusion, we could produce four FST-type proteins having different ligand selectivity in E. coli, and the results imply that economic production of a large amount of FST-type proteins with different ligand selectivity is possible to examine their potential use in meat-producing animals. Springer Berlin Heidelberg 2015-08-25 /pmc/articles/PMC4547976/ /pubmed/26302688 http://dx.doi.org/10.1186/s13568-015-0142-3 Text en © Lee et al. 2015 Open AccessThis article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made.
spellingShingle Original Article
Lee, Sang Beum
Park, Sung Kwon
Kim, Yong Soo
Production of bioactive chicken (Gallus gallus) follistatin-type proteins in E. coli
title Production of bioactive chicken (Gallus gallus) follistatin-type proteins in E. coli
title_full Production of bioactive chicken (Gallus gallus) follistatin-type proteins in E. coli
title_fullStr Production of bioactive chicken (Gallus gallus) follistatin-type proteins in E. coli
title_full_unstemmed Production of bioactive chicken (Gallus gallus) follistatin-type proteins in E. coli
title_short Production of bioactive chicken (Gallus gallus) follistatin-type proteins in E. coli
title_sort production of bioactive chicken (gallus gallus) follistatin-type proteins in e. coli
topic Original Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4547976/
https://www.ncbi.nlm.nih.gov/pubmed/26302688
http://dx.doi.org/10.1186/s13568-015-0142-3
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