Expression of active human GRO beta and GRO gamma neutrophil chemotactic proteins in E. coli.
Human GRO alpha, GRO beta, and GRO gamma are neutrophil chemoattractants structurally related to IL-8 and compete with IL-8 for binding to IL-8 receptors on neutrophils. These proteins are part of a large superfamily of chemotactic cytokines, the "chemokines," members of which share striking structural similarities. We have expressed GRO cDNA's in Escherichia coli as fusions to the MalE gene product, maltose-binding protein (MBP), in a way that allows separation of GRO and MBP moieties by factor Xa cleavage. GRO beta and GRO gamma expressed in bacteria were active in in vitro chemotaxis assays and were as effective as IL-8 in inducing chemotactic migration of neutrophils. Recombinant GRO beta was chemotactic rather than chemokinetic when tested by checkerboard analysis while GRO gamma showed evidence of chemokinetic as well as chemotactic activity. The activities of GRO beta and GRO gamma were not species-specific as both proteins were active on rat as well as human neutrophils and were inhibitable by antibodies raised against CINC, the rat GRO homolog. These data indicate that the MBP fusion protein expression system provides a rapid and simple method for obtaining large quantities of members of the chemokine protein family for biological uses.
Zagorski
J
J
Department of Health Sciences, Monsanto Company, St. Louis, Missouri 63167.
DeLarco
J E
JE
D018528:ATP-Binding Cassette Transporters; D001483:Base Sequence; D001681:Biological Assay; D002352:Carrier Proteins / Q000096:biosynthesis / Q000235:genetics / Q000494:pharmacology; D054360:Chemokine CXCL1; D019743:Chemokines, CXC; D002630:Chemotactic Factors / Q000096:biosynthesis* / Q000235:genetics / Q000494:pharmacology*; D002634:Chemotaxis, Leukocyte / Q000187:drug effects; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D004305:Dose-Response Relationship, Drug; D004926:Escherichia coli / Q000235:genetics; D029968:Escherichia coli Proteins; D006133:Growth Substances / Q000096:biosynthesis* / Q000235:genetics / Q000494:pharmacology*; D006801:Humans; D036341:Intercellular Signaling Peptides and Proteins; D058619:Maltose-Binding Proteins; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D009004:Monosaccharide Transport Proteins; D009504:Neutrophils / Q000187:drug effects; D033902:Periplasmic Binding Proteins; D016133:Polymerase Chain Reaction; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000494:pharmacology; D017421:Sequence Analysis
D018528:ATP-Binding Cassette Transporters; C475829:CXCL1 protein, human; D002352:Carrier Proteins; D054360:Chemokine CXCL1; D019743:Chemokines, CXC; D002630:Chemotactic Factors; C513297:Cxcl1 protein, rat; D018076:DNA, Complementary; D029968:Escherichia coli Proteins; D006133:Growth Substances; D036341:Intercellular Signaling Peptides and Proteins; C461619:MalE protein, E coli; D058619:Maltose-Binding Proteins; D009004:Monosaccharide Transport Proteins; D033902:Periplasmic Binding Proteins; D011993:Recombinant Fusion Proteins; C461599:maltose transport system, E coli