Strategies for expression and solubility analysis
Poster Nov 26, 2010
Production of soluble protein is one of the major bottlenecks that precede crystallographic studies.During the last years several techniques and strategies have been developed to address this problem. However, many of them imply an economical cost and technologies that are not always available. We will describe a general plan for protein solubility analysis by using a combination of four different but complementary strategies. In this plan, different constructs of a protein interest are designed and cloned in a library of expression vectors. All constructs are subsequently tested for bacterial overexpression using different strains at different culture conditions. The last step consists of a solubility screening for each construct tested for a specific strain and expression condition, by using different buffers designed according to a semi-rational approach.
These results support the use of hSC-CM and MEA technology for preclinical assessment of proarrhythmic risk within the proposed CiPA paradigm, and, more generally, demonstrate that automation of the CM-MEA assay can achieve high reliability and throughput for cardiac risk assessment in vitro.READ MORE