November 1, 2014 (Vol. 34, No. 19)

Gail Dutton

E. coli Strains Sustain Overexpression and Avoid Protein Aggregation, Toxicity, and Misfolding

Xbrane Bioscience has developed a protein expression system for Escherichia coli that eliminates much of the trial and error associated with E. coli strains, which typically offer little control over expression intensity. By designing the expression system to fit the protein, the company’s founders at Stockholm University’s Center for Biomembrane Research (CBR) were able to increase expression, yield, and quality of the produced protein.

The key is a modular approach that tunes each parameter for the protein of interest, drawing on Xbrane’s extensive strain library and customized research. That approach evolved from initial research in membrane protein overexpression work and, in 2008, formed the basis of Xbrane Bioscience.

At the time, expression systems offered either high or very low expression intensity, recalls Siavash Bashiri, CEO. Consequently, companies had to balance the need for high expression levels against the problems of protein aggregation, toxicity, and misfolding. “There was nothing in between,” he says.

Since then, however, Xbrane, with the input of its founders and other academic and industry partnerships, has developed expression systems that Bashiri says can resolve those challenges. Moreover, Xbrane can tailor these systems to suit individual customers. Xbrane, asserts Bashiri, offers strain development and protein production services that are built around whatever protein the customer desires: “Rather than search thousands of production strains, our approach helps customers quickly find the most optimal settings for expression of the target.”

Taking Control

Xbranes’s expression systems include Lemo, an E. coli BL21(DE3) strain engineered to overexpress protein, and Rhamex, a rhamnose-inducable protein production technology. Either system may be customized via OptiXpress, an Xbrane service that systematically optimizes strain and production parameters.

“Lemo is a T7-based system that works with many popular E. coli expression strains,” Bashiri explains. It optimizes overexpression of any protein using one, rather than many, strains. “With Lemo, you can titrate to set expression levels. This acts like a brake to the system and avoids the problems of aggregation, toxicity, and folding, and you get higher yields and more soluble protein.” It is compatible with all T7-based expression systems such as the well-known pET system.

In an optimization experiment, Xbrane increased expression between 10- and 30-fold using E. coli BL21(DE3) cells and the pET expression system. These gains were made by tuning parameters such as growth medium; induction strength and time; and growth and induction temperature. Other parameters may be adjusted. “One of the most important parameters is the tuning of the expression intensity,” Bashiri notes.

Rhamex, the second system to be commercialized by Xbrane, is E. coli strain independent. It can be used to increase expression even in toxic or otherwise challenging E. coli strains.

The benefits, according to Xbrane literature, include the “ability to express larger amounts of host-toxic and properly folded (active) proteins than other systems.” Because induction is slower using this system and can be precisely tuned, its protein production is highly uniform. Also, according to Bashiri, its expression levels can equal those of the T7 expression system.

In addition to producing proteins, Xbrane also is developing its optiXpress service model. For instance, “Biopharmaceutical, diagnostics, and medical device companies needing to increase expression or yields in E. coli can come to us for help with their internal projects,” Bashiri points out. Xbrane then uses its systems in combination with its in-house strain library to produce optimized, customized E. coli strains for protein production. The company also licenses its technology and works collaboratively with partners to develop future products.

Xbrane’s platforms include Lemo, an engineered E. coli strain, and Rhamex, a rhamnose-induceable system. Xbrane also offers OptiXpress, a service that emphasizes a target-centric approach to optimization.

Partnering for Growth

“Our expertise is in protein expression,” Bashiri emphasizes. “Ultimately, we want to be a preferred partner in developing expression systems; nearer-term, we aim to produce our own proteins off the shelf. We also might look at partnering for biosimilars.”

Reaching those goals involves outsourcing key competencies throughout the company. As a small company, Xbrane needs partners with proven track records in a broad array of fields.

Xbrane’s first commercial partnership began in 2009, when it collaborated with New England Biolabs to market Lemo. “We also are looking to different partners in the services industry,” Bashiri adds. Potential partners with expertise in protein purification and other downstream processes, manufacturing, and services are of interest.

Xbrane’s staff continues to work with the company’s founders at Stockholm University. Besides availing itself of scientific input, Xbrane enjoys the financial support of venture capital investor Serendipity Innovations as well as the advice of outside experts in financial management, marketing, and other fields. Currently, the company’s growth is mainly in Europe. Nonetheless, as Bashiri indicates, Xbrane does “have customers globally for optiXpress.”

Xbrane Bioscience

Location: Stureplan 15, 111 45 Stockholm, Sweden
Phone: +46 734 34 3619
Principal: Siavash Bashiri, CEO
Number of Employees: 5
Focus: Xbrane Biosciences aims to simplify and expedite the production of recombinant proteins in Escherichia coli. The company develops and optimizes strains and production systems that accommodate individual targets, tuning expression intensities, and sustaining high yields.