Recombinant Protein Expression

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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

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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
Kinetics and affinity analysis	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.

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