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Recombinant Protein Expression

VectorBuilder offers customized recombinant protein production services, employing multiple expression platforms suitable for various applications, such as protein structure analysis, enzymatic assay development, drug discovery, diagnostics, and bioengineering. Our protein production process is highly optimized for generating high-quality recombinant proteins utilizing our custom recombinant protein expression vectors. Additionally, we can produce recombinant proteins from customer-supplied vectors.

Highlights

  • One-stop solution: from vector design all the way through to production-scale protein purification
  • Comprehensive platform: flexibility to select from various expression systems
  • Customization: freedom to add a variety of purification or detection tags on our highly intuitive vector design platform
  • High performance: a broad range of purification methods and QC services are available to ensure high quality

Recombinant Protein Expression Systems

Workflow for Protein Production

Recombinant protein expression service

FAQ

Which recombinant protein expression system should I choose?

All recombinant protein expression systems have their pros and cons which should be taken into consideration while selecting the optimal system for your project. The table below summarizes the pros and cons of each system.

Recombinant protein expression system Pros Cons
Bacteria
  • Cost-effective
  • Short production time
  • Technically simple
  • Can be scaled up easily
  • High protein yield
  • Proteins lack post-translational modifications
  • Codon usage is different from that of eukaryotes
  • Difficult to express certain proteins due to the accumulation of inclusion bodies
  • Some proteins are toxic and can inhibit bacterial growth
Mammalian cells
  • Produces proteins in their most natural state with all necessary post-translational modifications and proper folding
  • Suitable for secretory and membrane proteins
  • Most appropriate for producing therapeutic proteins
  • Long production time
  • Requires complex growth conditions
  • Scaling up can be difficult
  • Not suitable for intracellular proteins due to low yield
Insect
  • Performs majority of complex post-translational modifications and protein folding
  • Suitable for secretory, intracellular, and membrane proteins
  • Can be used for producing large protein complexes
  • High levels of protein expression compared to other eukaryotic expression systems
  • Highly scalable due to high-density, suspension cell cultures
  • Long production time
  • Requires complex growth conditions
  • Technically challenging
Cell-free system
  • Time-efficient synthesis completed in 3 hours
  • No need for live cells in the production of toxic, complex, or unstable proteins
  • Suitable for high-throughput protein expression and screening
  • Easy procedure compatible with automated processes
  • Easy to optimize the conditions
  • Limited modifications are available
Which protein tag should I use?

Tags are frequently utilized for recombinant protein production. They streamline the purification process, and certain tags have demonstrated improvement in protein solubility, yield, or purity. If the tag is attached to the protease cleavage site, it can be removed after purification, and the efficiency of cleavage varies depending on the target protein. The careful selection of an appropriate tag is crucial for downstream protein expression and purification. The table below provides an overview of commonly used tags along with their advantages and limitations.

Tag Commonly applied system Advantages Limitations
GST Bacteria, insect
  • Largely preserves the native structure of the protein
  • Enables protein purification under mild conditions
  • Easy cleavage
  • Enhances the solubility and expression of the protein
  • Large tag size
  • Dimerization may impact the target protein
  • Not suitable for purifying proteins under denaturing conditions
His All
  • Small tag size, therefore less impact on the protein
  • Low cost for metal-affinity chromatography
  • Low immunogenicity
  • Suitable for purifying proteins under denaturing conditions
  • Other endogenous metal-binding proteins in microbial hosts may be co-purified, therefore optimization is usually required
  • Does not facilitate protein folding and solubility
SUMO Bacteria, insect
  • Facilitates protein folding
  • Enhances the solubility and expression of the protein
  • May undergo non-specific cleavage by other bacterial proteins
  • Not suitable for purifying proteins under denaturing conditions
Flag Mammalian cells, insect
  • Small tag size, therefore less impact on the protein
  • Easy detection
  • Low non-specific binding
  • Poor efficacy for purification
MBP Bacteria, insect
  • Enhances the solubility and expression of the protein
  • Large tag size
  • Not suitable for purifying proteins under denaturing conditions
Fc Mammalian cells, insect
  • Enhances the solubility and expression of proteins
  • Suitable for secreted protein
  • Large tag size

For more information regarding tags, please visit protein tags.

What QC services does VectorBuilder offer?

All vectors cloned by VectorBuilder come with a 100% sequence guarantee. All recombinant proteins produced by VectorBuilder undergo stringent quality control. Default QC for most systems includes 1) validation of the protein expression vector by restriction digestion analysis and Sanger sequencing; and 2) determination of protein concentration and purity by A260/280 measurement and SDS-PAGE. Common additional QC services are shown in the table below, which can be provided upon request.

Additional QC services Method
Endotoxin test LAL
Protein characterization Western Blot
Intact MS (reduced)
SEC-HPLC
SEC-MALs
Protein N-terminal sequencing
Host cell protein test
Tag removal Protease digestion
Kinetics and affinity analysis Octet
Biacore
Pathogen testing panel for rodents

                                                 Please inquire about other QC services.

What are the available purification methods?

VectorBuilder offers a wide range of protein purification methods, including size exclusion chromatography (gel filtration), affinity purification, ion exchange chromatography, and hydrophobic chromatography. For His-tagged or GST-tagged recombinant proteins, affinity chromatography is typically included, while for tag-free recombinant proteins, size exclusion chromatography (gel filtration) is typically included. The selection of the purification method(s) will be determined based on the construct design and protein characteristics.