HETEROLOGOUS EXPRESSION AND PURIFICATION OF NisA, THE PRECURSOR PEPTIDE OF LANTIBIOTIC Nisin FROM LACTOCOCCUS LACTIS
ACTA BIOLOGICA HUNGARICA, cilt.63, sa.2, ss.301-310, 2012 (SCI-Expanded, Scopus)
- Yayın Türü: Makale / Tam Makale
- Cilt numarası: 63 Sayı: 2
- Basım Tarihi: 2012
- Doi Numarası: 10.1556/abiol.63.2012.2.11
- Dergi Adı: ACTA BIOLOGICA HUNGARICA
- Derginin Tarandığı İndeksler: Science Citation Index Expanded (SCI-EXPANDED), Scopus
- Sayfa Sayıları: ss.301-310
- Bursa Uludağ Üniversitesi Adresli: Evet
Özet
The lantibiotic nisin is a ribosomally synthesised and post-translationally modified antimicrobial peptide produced by strains of Lactococcus lactis, and used as safe and natural preservative in food industry. The nisA structural gene encodes ribosomally synthesised and biologically inactive a 57 amino acid precursor peptide (NisA) which undergoes several post-translational modifications. In this study, we report the expression of precursor nisin as a His6-tagged peptide in Escherichia coli and its purification using a nickel affinity column. The technique of spliced-overlap extension PCR was used to amplify the nisA gene and the T7 promoter region of pET-15b vector. This approach was used to introduce six histidine residues at the C-terminus of prenisin. The identity of the expressed peptide was confirmed by N-terminal sequencing. The expressed His-tagged prenisin was purified under denaturing conditions, and named as prenisin-His6. The purified prenisin-His6 was analyzed by SDS-PAGE, Western blotting and mass spectroscopy. These results showed that the nisin precursor peptide can be successfully produced using an E. coli expression system.