Bacterial Expression of the scFv Fragment of a Recombinant Antibody Specific for Burkholderia pseudomallei Exotoxin

Yu Ching Su, Kue Peng Lim, Sheila Nathan

Research output: Contribution to journalArticle

13 Citations (Scopus)

Abstract

The scFv antibody towards the Burkholderia pseudomallei exotoxin was previously constructed by phage display and exhibited good specificity towards the exotoxin. We report here the optimization of the scFv expression in an E. coli expression system. Four different E. coli strains (ER2537, TG1, HB2151, and XL1-Blue) were examined for optimal expression of the scFv protein. Two types of carbon source (i.e. 0.2% glucose and 0.2% glycerol) were also tested for their ability to induce the scFv expression. Cells that carried the scFv construct were grown at 30°C and induced with 0.05 mM IPTG. The expression was then monitored by SDS-PAGE, Western blotting, and indirect ELISA. The Western blot profile showed different levels of the scFv expression among the host strains; XL1-Blue exhibited the highest level of the scFv protein expression. Glycerol at a concentration of 0.2% (v/v) significantly increased the scFv protein expression level when compared to 0.2% (w/v) glucose. Further optimization demonstrated that the scFv protein expression in XL1-Blue was the most optimal with a glycerol concentration as low as 0.05%. However, by indirect ELISA, only the scFv protein that was expressed in 0.2% (v/v) glycerol exhibited high specificity towards the Burkholderia pseudomallei exotoxin.

Original languageEnglish
Pages (from-to)493-498
Number of pages6
JournalJournal of Biochemistry and Molecular Biology
Volume36
Issue number5
Publication statusPublished - 30 Sep 2003

Fingerprint

Burkholderia pseudomallei
Exotoxins
Glycerol
Antibodies
Proteins
Escherichia coli
Western Blotting
Enzyme-Linked Immunosorbent Assay
Isopropyl Thiogalactoside
Glucose
Single-Chain Antibodies
Bacteriophages
Polyacrylamide Gel Electrophoresis
Carbon
Display devices

Keywords

  • Antibody
  • E. coli
  • Exotoxin
  • Protein expression
  • ScFv

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology

Cite this

Bacterial Expression of the scFv Fragment of a Recombinant Antibody Specific for Burkholderia pseudomallei Exotoxin. / Su, Yu Ching; Lim, Kue Peng; Nathan, Sheila.

In: Journal of Biochemistry and Molecular Biology, Vol. 36, No. 5, 30.09.2003, p. 493-498.

Research output: Contribution to journalArticle

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