Maltose binding protein-fusion enhances the bioactivity of truncated forms of pig myostatin propeptide produced in E. coli

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Abstract

© 2017 Lee et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Myostatin (MSTN) is a potent negative regulator of skeletal muscle growth. MSTN propeptide (MSTNpro) inhibits MSTN binding to its receptor through complex formation with MSTN, implying that MSTNpro can be a useful agent to improve skeletal muscle growth in meat-producing animals. Four different truncated forms of pig MSTNpro containing N-terminal maltose binding protein (MBP) as a fusion partner were expressed in E. coli, and purified by the combination of affinity chromatography and gel filtration. The MSTN-inhibitory capacities of these proteins were examined in an in vitro gene reporter assay. A MBP-fused, truncated MSTNpro containing residues 42±175 (MBP-Pro42-175) exhibited the same MSTNinhibitory potency as the full sequence MSTNpro. Truncated MSTNpro proteins containing either residues 42±115 (MBP-Pro42-115) or 42±98 (MBP-Pro42-98) also exhibited MSTNinhibitory capacity even though the potencies were significantly lower than that of full sequence MSTNpro. In pull-down assays, MBP-Pro42-175, MBP-Pro42-115, and MBPPro42- 98 demonstrated their binding to MSTN. MBP was removed from the truncated MSTNpro proteins by incubation with factor Xa to examine the potential role of MBP on MSTN-inhibitory capacity of those proteins. Removal of MBP from MBP-Pro42-175 and MBP-Pro42-98 resulted in 20-fold decrease in MSTN-inhibitory capacity of Pro42-175 and abolition of MSTN-inhibitory capacity of Pro42-98, indicating that MBP as fusion partner enhanced the MSTN-inhibitory capacity of those truncated MSTNpro proteins. In summary, this study shows that MBP is a very useful fusion partner in enhancing MSTN-inhibitory potency of truncated forms of MSTNpro proteins, and MBP-fused pig MSTNpro consisting of amino acid residues 42±175 is sufficient to maintain the full MSTN-inhibitory capacity.

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Lee, S. B., Park, S. K., & Kim, Y. S. (2017). Maltose binding protein-fusion enhances the bioactivity of truncated forms of pig myostatin propeptide produced in E. coli. PLoS ONE, 12(4). https://doi.org/10.1371/journal.pone.0174956

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