High-Yield Production of Bioactive Flatfish (Paralichthys olivaceus) Myostatin-1 Prodomain in Escherichia coli
Sang Beum Lee, Jeong Hwan Kim, Hyung-Joo Jin, Il Shik Shin and Yong Soo Kim
DOI : 10.3844/ajbbsp.2012.195.201
American Journal of Biochemistry and Biotechnology
Volume 8, Issue 3
Myostatin Propeptide (MstnPro), the N-terminal part of Mstn precursor molecule, binds to Mstn and suppress its activity. Mstn is a potent negative regulator of skeletal muscle growth in many animal species, thus MstnPro has a great potential to be used as an agent to enhance skeletal muscle growth in animal species. Bioactive fish Myostatin-1 Propeptide (Mstn1Pro) proteins were recently expressed in soluble forms in E. coli using Maltose Binding Protein (MBP) as a fusion partner. However, the yield of purified protein was too low to realize the benefit of large production capacity of the E. coli expression system. The objective of this study was to examine whether the yield of bioactive, recombinant flatfish (Paralichthys olivaceus) Mstn1Pro (poMstn1Pro) production can by improved through a proper selection of E. coli strain and other expression parameters. The poMstn1Pro gene construct, previously cloned into pMAL-c2 vector downstream of the Maltose-Binding Protein (MBP) gene, were transformed and expressed in E. coli K12BT1 strain. The MBP-poMstn1Pro protein was purified by the amylose-resin affinity chromatography. About 51 mg of soluble MBP-poMstn1Pro were purified by affinity chromatography from a liter culture. Pull-down assay and bioactivity test demonstrated the binding of MBP-poMstn1Pro to Mstn and its Mstn-suppressing capacity, indicating that the N-terminal fusion of MBP in the flatfish Mstn1Pro does not affect either the binding of flatfish Mstn1Pro to Mstn or its Mstn-suppressing activity. Results of this study demonstrate that a large production of MBP-poMstn1Pro is possible using an E. coli system to investigate the potential of MBP-poMstn1Pro to enhance muscle growth in aquaculture species or the binding characteristics of Mstn1Pro with its partners.
© 2012 Sang Beum Lee, Jeong Hwan Kim, Hyung-Joo Jin, Il Shik Shin and Yong Soo Kim. 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.