Preparing active growth factor dimers from inclusion bodies in high yield

摘要

Preparation of biologically active dimers (I) of recombinant human growth factors (II) of the cysteine knot family is new starting from cellular inclusion bodies (IB). IB are solubilized in an aqueous alkaline, urea-containing medium in the presence of a reducing agent, applied to an ion-exchange column, renatured, (e.g. refolded, dimerized) and re-oxidized aerobically, by washing with a decreasing urea gradient. The product is then eluted with a gradient of an inorganic salt, and the product is further purified by conventional methods of protein purification. Alternatively IB's are dissolved in aqueous medium containing solubilizing agent and reducing agent, are optionally gel filtered under denaturing conditions, then refolded to association-competent monomers in the presence of a redox agent (and optionally of solubilizing agent), by gel filtration. The product, present in the gel filtration eluate, is then converted to active dimer using a redox agent.