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   Protein Expression in E.coli and Pegylation Development

Recombinant Protein Expression in E.coli

E. coli is often the best expression system for many classes of therapeutic proteins, including cytokines, growth factors, enzymes, and nanobodies. It offers the advantages of quick turnover, easy scalability, and a high expression level. The service starts with the protein sequence provided by the client. We then test the suitability of expression methods (e.g., periplasmic secretion, intracellular soluble or inclusion body; tag or non-tag), expression and purification parameters, and finally scale up and perform product characterizations.

Since most of proteins expressed in E. coli will be in the form of inclusion body without biological activity, we use our proprietary protein refolding technology to develop a high-yield refolding procedure for the target protein. We develop a suitable purification procedure to purify the target protein, especially for therapeutic proteins. For example, although affinity tags are widely used for reagent proteins and offer an effective way to purify a protein of interest, including an affinity tag in a therapeutic protein may cause serious concerns in the regulatory approval process, and adding a tag cleavage step is often prohibitory in CMC development.

One of the disadvantages of an E. coli expression system is that the expressed proteins have no glycosylation, which often leads to short in vivo half-life. Pegylation is a proven method to effectively increase a protein’s half-life to make it comparable to or longer than that of CHO expressed glycosylated proteins, and also reduces its possible immunogenicity at the same time. Therefore, our service package includes a pegylation development option. We will use our Protein Mutation & Sequence Variant Analysis and Antibody & Protein Characterization platforms to analyze protein secondary structure and identify the optimal amino acid positions for specific PEG conjugation.

Process Flow Diagram

Please contact us for your protein expression inquiries or requests.

Application of protein expression service:

1. Expression of the recombinant proteins such as cytokines, interferons and other signaling molecules, enzymes, nanobodies, etc.

2. Development of therapeutic fusion proteins using our patented AT-fusion platform. This platform was developed to express human fusion protein in E. coli with high yield and high biological activity, as well as an excellent PK profile.

Clients Input: a protein or nanobody amino acid sequence.
Deliverable: expressed and purified protein at the amount requested by the client.

Related Service & Products

  1. Therapeutic Fusion Proteins & Nanobodies Development
  2. Protein Mutation & Sequence Variant Analysis
  3. Therapeutic antibody and Protein Characterizations