Protein Sciences

A: Absolutely. Besides His-tag, we also offer Flag-tag for miniproteins. But if your project is aiming for miniproteins with specific constructs, we can purify them without any tags, and then use conventional chromatography for purification.

A: We run SEC-HPLC to determine the purity of miniproteins. This technique separates the miniproteins based on their size, allowing us to pinpoint their relative abundance.

A: We’ve got various analytical methods to measure the stability of miniproteins. For instance, we can measure their melting temperature (TM) or examine their structure with circular dichroism (CD).

A: Our Protein Sciences (PS) Department at WuXi Biologics focuses on the research domain, with our expertise primarily rooted in smaller-scale projects. However, when scaling up to 1 L using our CHO transient platform, we can achieve yields of 400-500 mg of protein. We are capable of scaling up further without an upper limit.

A: We do not offer computational services for miniprotein property improvement. The field of miniproteins is rapidly advancing, and we lean on our client’s expertise to guide protein development.

A: After producing miniproteins, our go-to downstream assays include BLI, SPR, and occasionally ELISA for certain cases. These methods, especially BLI, are highly efficient for HTP screening of miniproteins within a short timeframe. Additionally, we can conduct sophisticated, low-throughput assays for your specialized proteins.

A: We consider expression level >100 mg/L to be typical for most miniproteins. In a case study where we expressed 48 miniproteins in CHO cells, the average yield was 197 mg/L.

A: Our understanding of sequence homology across various species and its impact on miniprotein expression is limited, since these proteins are designed de novo. Generally, we observe higher expression levels in CHO compared to E. coli, regardless of sequence homology. Other factors such as the maturity of the design algorithm and the structural characteristics of the miniproteins also influence expression levels.

A: The miniproteins’ structures are crucial for their yields. Miniproteins with stable hydrophobic cores with proper folding are more likely to be expressed successfully in both E. coli and CHO. For miniproteins designed to function intracellularly with free cysteines, E. coli is often preferred. Conversely, miniproteins lacking free cysteines or those with disulfide bonds are better expressed in CHO. Additionally, when miniproteins serve as building blocks for mAbs or bsAbs, CHO is recommended to achieve optimal expression levels.

A: Yes, we have encountered cases where miniproteins that are insoluble in E. coli were successfully expressed in a soluble form in CHO cells.

A: Our expression system produces miniproteins with lengths between 80 and 200 amino acids. The constraint on the maximum size is primarily related to the capabilities of algorithmic design rather than our expression system. Currently, we do not provide services for sequence design.

A: While the pI range of miniproteins typically does not pose issues, miniproteins with a high concentration of basic amino acids can lead to proteolysis in both E. coli and CHO cells. Monitoring the proteolysis profile of these miniproteins is essential for in vivo therapeutic applications to ensure their stability and effectiveness. To support this, we offer serum stability tests as part of our micro developability services.

A: Yes, we offer both in silico immunogenicity prediction to identify sequences with potential immunogenicity issues and ex vivo immunogenicity assays to assess the immunogenicity risk of miniproteins.