Modular Broad-Host-Range Expression Vectors for Single-Protein and Protein Complex Purification

Barna D. Fodor, Ákos T. Kovács, Róbert Csáki, E. Hunyadi-Gulyás, E. Klement, Gergely Maróti, Lívia S. Mészáros, Katalin F. Medzihradszky, G. Rákhely, K. Kovács

Research output: Contribution to journalArticle

29 Citations (Scopus)

Abstract

A set of modular broad-host-range expression vectors with various affinity tags (six-His-tag, FLAG-tag, Strep-tag II, T7-tag) was created. The complete nucleotide sequences of the vectors are known, and these small vectors can be mobilized by conjugation. They are useful in the purification of proteins and protein complexes from gram-negative bacterial species. The plasmids were easily customized for Thiocapsa roseopersicina, Rhodobacter capsulatus, and Methylococcus capsulatus by inserting an appropriate promoter. These examples demonstrate the versatility and flexibility of the vectors. The constructs harbor the T7 promoter for easy overproduction of the desired protein in an appropriate Escherichia coli host. The vectors were useful in purifying different proteins from T. roseopersicina. The FLAG-tag-Strep-tag II combination was utilized for isolation of the HynL-HypC2 protein complex involved in hydrogenase maturation. These tools should be useful for protein purification and for studying protein-protein interactions in a range of bacterial species.

Original languageEnglish
Pages (from-to)712-721
Number of pages10
JournalApplied and Environmental Microbiology
Volume70
Issue number2
DOIs
Publication statusPublished - Feb 2004

Fingerprint

Host Specificity
host range
purification
Trp-Ser- His-Pro-Gln-Phe-Glu-Lys
Thiocapsa roseopersicina
protein
Proteins
proteins
Methylococcus capsulatus
Rhodobacter capsulatus
promoter regions
protein-protein interactions
Hydrogenase
plasmids
plasmid
Escherichia coli
nucleotide sequences
maturation
harbor
Plasmids

ASJC Scopus subject areas

  • Environmental Science(all)
  • Biotechnology
  • Microbiology

Cite this

Modular Broad-Host-Range Expression Vectors for Single-Protein and Protein Complex Purification. / Fodor, Barna D.; Kovács, Ákos T.; Csáki, Róbert; Hunyadi-Gulyás, E.; Klement, E.; Maróti, Gergely; Mészáros, Lívia S.; Medzihradszky, Katalin F.; Rákhely, G.; Kovács, K.

In: Applied and Environmental Microbiology, Vol. 70, No. 2, 02.2004, p. 712-721.

Research output: Contribution to journalArticle

Fodor, Barna D. ; Kovács, Ákos T. ; Csáki, Róbert ; Hunyadi-Gulyás, E. ; Klement, E. ; Maróti, Gergely ; Mészáros, Lívia S. ; Medzihradszky, Katalin F. ; Rákhely, G. ; Kovács, K. / Modular Broad-Host-Range Expression Vectors for Single-Protein and Protein Complex Purification. In: Applied and Environmental Microbiology. 2004 ; Vol. 70, No. 2. pp. 712-721.
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