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Expression and characterization of antimicrobial peptide ABP-CM4 in methylotrophic yeast Pichia pastoris

Paper ID Volume ID Publish Year Pages File Format Full-Text
36155 45123 2006 6 PDF Available
Title
Expression and characterization of antimicrobial peptide ABP-CM4 in methylotrophic yeast Pichia pastoris
Abstract

Antibacterial peptides CM4 (ABP-CM4) is a linear cationic peptide that has antimicrobial properties. To explore a new approach of expression of ABP-CM4 in methylotrophic yeast Pichia pastoris, the gene of ABP-CM4 was obtained by recursive PCR (rPCR) and cloned into the vector pPICZaA. The SacI-linearized plasmid pPICZaA-CM4 was transformed into P. pastoris GS115 by electroporation. The expression was induced about 72 h with 0.5% methanol at 20 °C, supplied with 2% casamino acids to avoid proteolysis, and approximately 40 mg ABP-CM4 was secreted into 1 L of medium. Recombinant ABP-CM4 was purified through size-exclusion chromatography and 15 mg pure active ABP-CM4 was obtained from 1 L culture. Tricine–SDS–PAGE indicated that recombinant ABP-CM4 was secreted as a protein of around 3.8 kDa. The recombinant ABP-CM4 appears to be successfully expressed, as it displays antibacterial and antifungal activity (antibacterial assay, polyarylamide gel electrophoresis, and antifungal assay) indistinguishable from those of natural protein.

Keywords
Antibacterial peptide, ABP-CM4, Antifungal, Recursive PCR, pPICZaA, P. pastoris
First Page Preview
Expression and characterization of antimicrobial peptide ABP-CM4 in methylotrophic yeast Pichia pastoris
Publisher
Database: Elsevier - ScienceDirect
Journal: Process Biochemistry - Volume 41, Issue 2, February 2006, Pages 251–256
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering