Innovative Tools for Molecular and Cell Biology

Innovative Tools for Molecular and Cell Biology

Products

BRP Plasmids & Competent BRP Cells

For the Release of Cellular or Recombinant Proteins into the Culture Medium

 Cotransformation of recombinant plasmid and BRP vector (in one or two steps)

1.
Cotransformation
of recombinant plasmid and BRP vector (in one or two steps)

 Induction of BRP, which then activates Phospholipase A

2.
Induction of BRP, which then activates Phospholipase A

 Excretion of cytosolic and periplasmic proteins into the cell culture medium

3.
Excretion of cytosolic and periplasmic proteins into the cell culture medium

Features

  • Vectors expressing the Bacteriocin Release Protein (BRP)
  • Protein excretion into E. coli culture medium without the need of a signal sequence
  • Release of lethal proteins
  • Preventing protein degradation by cytoplasmic proteases
  • Avoiding inclusion bodies
  • Release of periplasmic proteins
  • Large-scale biotechnological production of proteins in a continuous culture
  • Simplified protein purification from culture medium
  • Vectors can be co-transformed with ColE1 vectors
  • Alternatively, we are offering competent BRP-transformed cells

Description

  • The plasmids pJL3 and pSW1 express the Bacteriocin Release Protein (BRP) upon induction, which initiates release of periplasmic and cytoplasmic E. coli proteins into the culture medium. Being compatible with most of the commonly used expression vector systems (e.g. ColE1 vectors like pAX, PheBo, pBR322 derivatives), BRP vectors can be co-transformed with the vector producing the recombinant protein of interest. Induction of the BR-Protein with IPTG (pJL3) or mitomycin C (pSW1) will cause an activation of phospholipase A in the outer E. coli membrane. This results in the formation of permeable zones in the cell membranes, through which proteins are released into the medium. A moderate induction prevents lysis of producer cells, also making the system suitable for large-scale protein production in a continuous culture. Since cloned proteins are no longer accumulated in the cytoplasm, problems associated with lethality of recombinant proteins, their preferential degradation or inclusion body formation are avoided. For your convenience, we also provide competent cells already transformed with one of the vectors.
Vector map of pJL3

Vector map of pJL3
ori, origin of replication; CmR, chloramphenicol resistance; lppp, E. coli lipoprotein promoter, lacpo, lac promoter operator system; lac I, lac repressor; BRP, Bacteriocin Release Protein.

 Vector map of pSW1

Vector map of pSW1
ori, origin of replication; TetR, tetracycline resistance; CmR, part of chloramphenicol resistance gene, not functional; pClo, pCloDF13 promoter; BRP, Bacteriocin Release Protein.

Examples

Proteins that have already been successfully released by activity of BRP from the E. coli periplasm:

ProteinSize in kDa
Bacillus penicillinase25
Aeromonas xylanase L135
Human IgG Fc region29
Human chimeric IgE/IgG Fc37
Human calcitonin 27
Guar alpha-galactosidase 40
Bacillus alpha-amylase 55
Bacillus cellulase (N-4 and 1139)58/92
Human growth hormone 21
Human tumor necrosis factor-alpha 17
beta-lactamase29
FaeE25

ORDER INFORMATION

  • For shipping and storage information please click on Order#.
Order# Description Amount Price Data Sheet
BRPJL3 BRP Plasmid, pJL3, lyophilized DNA 5 µg 283,00 PDF
BRPSW1 BRP Plasmid, pSW1, lyophilized DNA 5 µg 283,00 PDF
COMJL3 E.coli K12 transformed with pJL3 5x200 µl 356,00 PDF
COMSW1 E.coli K12 transformed with pSW1 5x200 µl 356,00 PDF
  • All prices are in EURO excl. VAT and shipping. For further pricing and order information please ask your local distributor.

Download

  • BRP Vectors and Competent Cells Handbook (PDF)