New fusion protein systems designed to give soluble expression in Escherichia coli

Three native E. coli proteins-NusA, GrpE, and bacterioferritin (BFR)-were s tudied in fusion proteins expressed in E. coli for their ability to confer solubility an a target insoluble protein at the C-terminus of the fusion pr otein. These three proteins were chosen based on their favorable cytoplasmi c solubility characteristics as predicted by a statistical solubility model for recombinant proteins in E. coli. Modeling predicted the probability of soluble fusion protein expression for the target insoluble protein human i nterleukin-3 (hIL-3) in the following order: NusA (most soluble), GrpE, BFR , and thioredoxin (least soluble). Expression experiments at 37 degrees C s howed that the NusA/hIL3 fusion protein was expressed almost completely in the soluble fraction, while GrpE/hIL-3 and BFR/hIL-3 exhibited partial solu bility at 37 degrees C. Thioredoxin/hIL-3 was expressed almost completely i n the insoluble fraction. Fusion proteins consisting of NusA and either bov ine growth hormone or human interferon-gamma were also expressed in E. coli at 37 degrees C and again showed that the fusion protein was almost comple tely soluble. Starting with the NusA/hIL-3 fusion protein with an N-termina l histidine tag, purified hIL-3 with full biological activity was obtained using immobilized metal affinity chromatography, factor Xa protease cleavag e, and anion exchange chromatography. (C) 1999 John Wiley & Sons, Inc.