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Expression and purification of a recombinant buforin derivative from Escherichia coli

Pyo, Sang-Hyun LU orcid ; Lee, Jae-Hyun ; Park, Heung-Bok ; Cho, Jin-Suk ; Kim, Hong-Rak ; Han, Byung-Hee and Park, Yeon-Sung (2004) In Process Biochemistry 39(11). p.1731-1736
Abstract
An Escherichia coli expression system was constructed for
production of a recombinant antimicrobial peptide buforin derivative
named buforin IIb and a large-scale process of cultivation and
purification was also developed. Inclusion bodies of fusion proteins
containing recombinant buforin IIb were released from
high-density-cultured cells by high-pressure homogenization. The
inclusion bodies solubilized in a buffer containing 8 M urea were incubated in cleavage buffer containing 1.7 M
hydroxylamine. The crude mixtures were fractionated by ammonium sulfate
precipitation and then washed with distilled water to remove salts. The
pellets were dissolved in 7 M urea solution, and treated with... (More)
An Escherichia coli expression system was constructed for
production of a recombinant antimicrobial peptide buforin derivative
named buforin IIb and a large-scale process of cultivation and
purification was also developed. Inclusion bodies of fusion proteins
containing recombinant buforin IIb were released from
high-density-cultured cells by high-pressure homogenization. The
inclusion bodies solubilized in a buffer containing 8 M urea were incubated in cleavage buffer containing 1.7 M
hydroxylamine. The crude mixtures were fractionated by ammonium sulfate
precipitation and then washed with distilled water to remove salts. The
pellets were dissolved in 7 M urea solution, and treated with
polyethyleneimine (PEI) to remove contaminated DNA. The recombinant
buforin IIb was purified by cation exchange chromatography and reverse
phase HPLC (RP-HPLC). The purity of the recombinant buforin IIb was
determined by RP-HPLC and SDS-PAGE and the molecular mass of the
recombinant buforin IIb analyzed by MALDI-TOF-MS showed good agreement
with the authentic buforin IIb. These results suggest that the
production method in this report is useful in obtaining a large quantity
of recombinant antimicrobial peptide. (Less)
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author
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publishing date
type
Contribution to journal
publication status
published
in
Process Biochemistry
volume
39
issue
11
pages
1731 - 1736
publisher
Elsevier
external identifiers
  • scopus:3042850735
ISSN
1359-5113
DOI
10.1016/j.procbio.2003.07.007
language
English
LU publication?
no
id
c8fb00a4-6ba6-4f84-bb51-8e356e8e8d2a
date added to LUP
2025-09-05 17:54:58
date last changed
2025-10-14 12:46:12
@article{c8fb00a4-6ba6-4f84-bb51-8e356e8e8d2a,
  abstract     = {{An <em>Escherichia coli</em> expression system was constructed for <br>
production of a recombinant antimicrobial peptide buforin derivative <br>
named buforin IIb and a large-scale process of cultivation and <br>
purification was also developed. Inclusion bodies of fusion proteins <br>
containing recombinant buforin IIb were released from <br>
high-density-cultured cells by high-pressure homogenization. The <br>
inclusion bodies solubilized in a buffer containing 8 M urea were incubated in cleavage buffer containing 1.7 M<br>
 hydroxylamine. The crude mixtures were fractionated by ammonium sulfate<br>
 precipitation and then washed with distilled water to remove salts. The<br>
 pellets were dissolved in 7 M urea solution, and treated with <br>
polyethyleneimine (PEI) to remove contaminated DNA. The recombinant <br>
buforin IIb was purified by cation exchange chromatography and reverse <br>
phase HPLC (RP-HPLC). The purity of the recombinant buforin IIb was <br>
determined by RP-HPLC and SDS-PAGE and the molecular mass of the <br>
recombinant buforin IIb analyzed by MALDI-TOF-MS showed good agreement <br>
with the authentic buforin IIb. These results suggest that the <br>
production method in this report is useful in obtaining a large quantity<br>
 of recombinant antimicrobial peptide.}},
  author       = {{Pyo, Sang-Hyun and Lee, Jae-Hyun and Park, Heung-Bok and Cho, Jin-Suk and Kim, Hong-Rak and Han, Byung-Hee and Park, Yeon-Sung}},
  issn         = {{1359-5113}},
  language     = {{eng}},
  month        = {{07}},
  number       = {{11}},
  pages        = {{1731--1736}},
  publisher    = {{Elsevier}},
  series       = {{Process Biochemistry}},
  title        = {{Expression and purification of a recombinant buforin derivative from <i>Escherichia coli</i>}},
  url          = {{http://dx.doi.org/10.1016/j.procbio.2003.07.007}},
  doi          = {{10.1016/j.procbio.2003.07.007}},
  volume       = {{39}},
  year         = {{2004}},
}