Simple purification of human antimicrobial peptide dermcidin (MDCD-1L) by intein-mediated expression in E.coli.
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.
Hong
Inpyo
I
Department of Bioengineering and Technology, Kangwon National University, Chuncheon 200-701, Korea.
Kim
Yong-Seok
YS
Choi
Shin-Geon
SG
D000900:Anti-Bacterial Agents / Q000302:isolation & purification* / Q000378:metabolism / Q000494:pharmacology; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D015870:Gene Expression; D006801:Humans; D047668:Inteins; D008837:Micrococcus / Q000187:drug effects; D010455:Peptides / Q000235:genetics* / Q000302:isolation & purification* / Q000378:metabolism / Q000494:pharmacology; D015202:Protein Engineering; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism / Q000494:pharmacology
D000900:Anti-Bacterial Agents; D010455:Peptides; D011993:Recombinant Fusion Proteins; C442243:dermcidin