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KMID : 0545120100200020350
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Journal of Microbiology and Biotechnology
2010 Volume.20 No. 2 p.350 ~ p.355
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Simple purification of human antimicrobial peptide dermcidin (MDCD-1L) by intein-mediated expression in E.coli
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Hong In-Pyo
Kim Yong-Seok
Choi Shin-Geon
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Abstract
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Among human antimicrobial peptides (hAMPs), DCD-1L has a broad spectrum of antimicrobial activity over a wide pH range and in high salt concentrations. It offers a promising alternative to conventional antibiotics. The 458-bp-long dermcidin cDNA was amplified by PCR using a human fetal cDNA library as a template. The 147-bp fragment of the MDCD-1L gene encoding an additional methionine residue was subcloned into the pTYB11 vector. Recombinant MDCD-1L was expressed as an intein fusion protein in E. coli, and then purified by affinity chromatography using chitin beads. A small peptide with a molecular mass of about 5 kDa was detected by tricine gel electrophoresis. The recombinant MDCD-1L peptide was purified from the gel and its amino acid sequence was determined by nanoLC-ESI-MS/MS analysis. The initiating amino acid, methionine, remained attached to the N-terminal region of recombinant MDCD-1L. Purified MDCD-1L showed antimicrobial activity against a Micrococcus luteus test strain.
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KEYWORD
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Antimicrobial peptides, DCD-1L, Dermcidin, Intein, Protein Expression
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