E. coli expression system

Despite the development of the eukaryotic expression systems, E. coli remains the most widely used expression system for recombinant proteins. Advantages of E. coli expression are low cost, high expression level, easy to scale up, short turnaround time. One point to keep in mind is that expression of foreign proteins in E. coli may result in the formation of protein aggregates, commonly known as inclusion bodies, as a result of imbalance between protein aggregation and solubilization. In this case, our protein refolding system are set to solubilize and refold the inclusion bodies and thereby obtain soluble, bioactive recombinant proteins.

Novoprotein’s bacteria expression system serves you with these advantages:

  • High Efficiency:

    Collection of more than 10 host strains and various proprietary or non-proprietary vectors guarantees the selection of the most efficient expression system.

  • High Productivity:

    Enhanced fusion tag technology and proprietary AIE technology are available in house to increase protein solubility and production, and to decrease purification cost, even for toxic proteins.

  • Advanced refolding:

    We have established and optimized over the years a sophisticated protein refolding system composed of 20 refolding buffers to increase the chance of obtaining soluble and bioactive proteins.

  • Large-Scale Production:

    We are able to produce up to 500L of E. coli cell culture for gram scale production.

  • Modifications:

    Metabolic labeling with stable isotopes (13C and 15N) and biotinylation are available.

  • Competitive Pricing:

    5mg protein at 80% purity from only $1000!

  • Procedure details

  • Timeframe and pricing

  • Case study

We start with your plasmid,cDNA,or simply the gene sequence information
We optimize the codon to maxmize the expression level in you choice of system
We test the expression in various vectors and E. coli strains, and then choose the most efficient expression vector and strain for your gene
Our 20 propriety reflolding buffers and 17 solubilization buffers ensure the highest protein productivity and quality
Our highly advanced chromatography skills and complex column system guarantee the purity as we promise
  • Your gene of interest
  • Evaluate and choose the suitable expression system
  • Codon Optimization
  • Sub-clone into E. coli Expression Vectors
  • Transfect expression plasmid into E. coli
  • Select high-expression transformants
  • Expression evaluation and optimization
  • Protein Refolding
  • Large scale expression
    (up to 500L,gram scale)
  • SDS Page Gel
  • Protein purification
    • Affinity column
    • Ion exchange chromatography
    • Hydrophobic chomatography
    • Size exclusion chromatography
  • Ready to go!
Gene Cloning and Expression Test
Strategy design Free 1 day
Codon optimization Free 2-3 weeks
Gene synthesis $0.40/bp
Subcloning gene of interest into expression vector $195.00
Expression strain construction and expression optimization $100.00 2 weeks
Pilot Scale Expression and Purification
2L bacterial cell culture From $1,000.00 2-4 weeks
5mg of protein (soluble, >80% purity)
Large Scale Production
Large-scale fermentation up to 500L with 60-80g cell/L From $1,000.00 1-2 weeks
Purification to gram-scale protein  
Protein production to kilogram scale Please inquire  
Additional Services
Protein refolding    
Endotoxin removal    
Metabolic labeling    
Biotinylation    
Lyophilization    
Process development    
   
  • Protein property:

    • Secreted cytokine
    • Expressed as pro-peptide (33kD), processed to mature form (14kD)
  • Challenges:

    Both pro-peptide and mature forms are expressed in inclusion body (IB)

  • Strategy:

    • Pro-peptide modification to facilitate the refolding
    • Solubilize and refold pro-peptide
    • Remove pro-peptide sequence
  • Deliverables:

    Refolded bioactive mature peptide above 95% purity

  • 1.Expression strain screening

    Fig.1 SDS-PAGE analysis of pro-peptide expression.
    Lane2: After inducing
    Lane3: Before inducing
  • 2.Inclusion body solubilization and pro-peptide refolding

    Fig.2 Solubilization and refolding of inclusion body in NP proprietary buffers.
    IB: Inclusion body
    R: Soluble and folded pro-peptide, Reduced
    NR: Soluble and folded pro-peptide, Non-Reduced
  • 3.Purification of refolded pro-peptide

  • Fig.3 IEX purification (SP-HP) of refolded pro-peptide
    S: Sample
    FT: Flow Through
    E1: Elution1
    E2: Elution2
  • 4.Mature peptide processing

    Fig.4 Pro-peptide processed into mature form with NP proprietary reagents and buffer.
    pp: Pro-peptide
    mp(R): Mature peptide (reduced)
    mp(NR): Mature peptide (non-reduced)
  • 5.Final product: refolded bioactive mature peptide with purity above 95% (Mass Spec confirmed)

    Fig.5 Final product analyzed by SDS-PAGE.
    Fraction1: 0.24mg/ml(A280)
    Fraction2: 0.42mg/ml(A280)
    Fraction3: 0.86mg/ml(A280)

For more information about our E. coli expression service, contact us at sales@novoprotein.com or visit our Service page. In addition, we also offer other protein expression services including yeast protein expression services, baculovirus expression services,mammalian expression service. To know how other scientists speak of us, check out our References.

YOUR CONCERNS

Will I get what I am promised?
Our counter-contract ensures your terms of expectation are met.

I feel like losing control of data generation!
Our service team keeps tight monitoring and tracking over each step where data are generated.