SYNTHESIS AND EXPRESSION OF A GENE ENCODING CIRCULAR ENTEROCIN AS-48 IN ESCHERICHIA COLI

  • Trần Ngọc Tân

Abstract

Enterocin AS-48 is a 70 amino acid circular peptide produced by Enterococcus faecalis and it shows broad spectrum of anti-microbial activity against Gram-positive and Gram-negative bacteria. In order to produce the peptide, the as-48 gene sequence was designed from GenBank and the 210 base pairs gene was artificially synthesized in two steps peR. In the step 1, two long primers with over-lapping sequences served as DNA template and in the step 2, two short primers were used to amplifY as-48 gene by PCR. The PCR product was cloned into pJETI vector and sequencing confirmed presence of 210 nucleotides as expected. To produce the recombinant mature enterocin AS-48 pepdide, the synthetic gene was cloned to the expression vector pTWIN under the control of T7 RNA promoter in E. coli ER2566. The recombinant AS-48 peptide was expressed as a fusion protein with 2 intein on either side (N and C terminal). The fusion protein was 63 kOa on SOS-PAGE and reacted with chitin antibody in Western blot, thus confirming its identify. This paper described the first design of synthetic gene as-48 and the fusion enterocin AS-48 peptide with promising antibacterial properties on agar plates.

Tác giả

Trần Ngọc Tân
điểm /   đánh giá
Published
2011-11-25
Section
Articles