Chemokines derived from soluble fusion proteins expressed in Escherichia coli are biologically active.
Chemokines are a class of low molecular weight proteins that are involved in leukocytes trafficking. Due to their involvement in recruiting immune cells to sites of inflammation, chemokines, and chemokine receptors have become an attractive class of therapeutic targets. However, when expressed in Escherichia coli chemokines are poorly soluble and accumulate in inclusion bodies. Several purification methods have been described but involve time-consuming refolding, buffer exchange, and purification steps that complicate expression of these proteins. Here, we describe a simple and reliable method to express chemokines as fusions to the protein NusA. The fusion proteins were largely found in the soluble fraction and could be readily purified in a single step. Proteolytic cleavage was used to obtain soluble recombinant chemokines that were found to be very active in a novel in vitro chemotaxis assays. This method could be applied to several alpha and beta human chemokines, suggesting that it is generally applicable to this class of proteins.
Magistrelli
Giovanni
G
NovImmune SA, 64 avenue de la Roseraie, 1211 Geneva 4, Switzerland.
Gueneau
Franck
F
Muslmani
Machadiya
M
Ravn
Ulla
U
Kosco-Vilbois
Marie
M
Fischer
Nicolas
N
D000818:Animals; D045744:Cell Line, Tumor; D018925:Chemokines / Q000032:analysis / Q000096:biosynthesis* / Q000737:chemistry* / Q000235:genetics; D003001:Cloning, Molecular / Q000379:methods; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism*; D008223:Lymphoma / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D015202:Protein Engineering; D011993:Recombinant Fusion Proteins / Q000032:analysis / Q000096:biosynthesis* / Q000737:chemistry*; D012995:Solubility
D018925:Chemokines; D011993:Recombinant Fusion Proteins