Expression and Identification of Apidaecin in Pichia Pastoris.

doi:10.13701/j.cnki.kqyxyj.2017.05.002

Abstract

Abstract: Objective: To express biologically active Apidaecin in pichia pastoris (P. pastoris) and detect its antibacterial activity. Methods: The EcoR I restriction sites and GST tag which was connected with DDDDK enterokinase site were inserted to the N-terminus of the Apidaecin gene by polymerase chain reaction (PCR). A termination codon and Not I restriction sites were inserted at the C-terminus. The amplified fragments were connected to the expression vector pPICZαA and the recombinant expression vector pPICZαA-GST-Apidaecin was established and the sequence was identified. The pPICZαA-GST-Apidaecin was electrotransfected into P.pastoris X33 and was fermented. The expression products were purified and the molecular weight was appraised by SDS-PAGE gel electrophoresis. The antibacterial activity was tested after the GST was excised by EK kinase. Results: SDS-PAGE gel electrophoresis showed that the molecular weight of the product was about 28KD which was identical with the theoretical molecular weight. After removal of GST fusion tag by enterokinase, the antibacterial activity of Apidaecin to E.coli BL21 was significant. Conclusion: The antimicrobial peptides of Apidaecin typecan successfully expressed in P. pastoris and had significant antibacterial activity.

CLC Number:

R780.1

SHI Wen, MA Rui, ZHOU Jian-ye, CHEN Li-ya, MA Yuan-yuan, HUANG Hui-min, ZHANG xiao-feng, YI Geng-yun, LI Zhi-qiang.. Expression and Identification of Apidaecin in Pichia Pastoris.[J]. Journal of Oral Science Research, 2017, 33(5): 471-474.

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