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Tropical Journal of Pharmaceutical Research
Pharmacotherapy Group, Faculty of Pharmacy, University of Benin, Benin City, Nigeria
ISSN: 1596-5996 EISSN: 1596-5996
Vol. 16, No. 4, 2017, pp. 811-818
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Bioline Code: pr17103
Full paper language: English
Document type: Research Article
Document available free of charge
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Tropical Journal of Pharmaceutical Research, Vol. 16, No. 4, 2017, pp. 811-818
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Simple and efficient expression of codon-optimized mouse leukemia inhibitory factor in Escherichia coli
Niu, Dewei; Wu, Qiuhong; Huang, Luyuan; Yang, Enze; Huang, Meiyan; Liu, Yong & Wang, Feng
Abstract
Purpose: To obtain a higher yield of mouse leukemia inhibitory factor to maintain the proliferation
potential of pluripotent stem cells at a low cost.
Methods: A method was designed to produce recombinant mLIF protein (rmLIF) in Escherichia coli.
Through analysis of rmLIF sequence, it was found that rare codons were interspersed. After mutation
from rare codons to Escherichia coli (E. coli) preferred ones were selected, the mutated gene mLIFm
was cloned into pET15b vector. The pET15b-mLIFm was then transformed into Rosetta-gami strain and
induced with optimal conditions at 18 oC for 16 h. Mass spectrometry was carried out to identify the
peptides.
Results: After purification, the yield of the codon-optimized rmLIFm was 141 mg/L, compared with 110
mg/L for the original rmLIF. Mass spectral analysis showed the presence of four major peptides each
with an intensity > 10 % at m/z 1031.57, 1539.82, 1412.01 and 2229.10 in mLIFm, respectively. His-tagged
rmLIFm fusion protein displayed the potential to maintain the morphology of undifferentiated
mouse embryonic stem cells (mESCs), which were positive for mESCs markers (Oct-4, Nanog, Sox-2,
stage-specific embryonic antigen-1).
Conclusion: The findings provide a means to produce mLIF in a short, useful, cost-effective and
environmentally-friendly manner, and thus lays a foundation for further studies of mLIF.
Keywords
Leukemia inhibitory factor, Mutated gene, Protein expression, Purification, Stem cells, Peptides, Escherichia coli
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