title: Streptococcal protein G, expressed by streptococci or by Escherichia coli, has separate binding sites for human albumin and IgG.
abstract: Protein G is expressed at the cell surface of certain group C and group G streptococcal strains. The protein shows a unique and specific affinity for the Fc region of mammalian polyclonal and monoclonal immunoglobulin G (IgG). We have cloned the streptococcal gene coding for protein G into E. coli, using phage lambda as the vector. The protein G produced by E. coli infected with this phage was detected and analysed in Western blot experiments using radiolabelled IgG Fc fragments as a probe. Three major IgG Fc-binding bands were obtained corresponding to apparent mol. wts of 47,000, 57,000 and 65,000, respectively. Analysis of the expression in E. coli indicates that this heterogeneity is caused by a post-translational degradation of the molecule before lysis of the lambda infected E. coli cells occurred. The protein G produced in E. coli was purified by affinity chromatography on IgG-Sepharose followed by gel-filtration on Sephadex G-200. This highly purified E. coli-produced protein G was compared to protein G solubilized by papain from streptococci, in direct binding experiments and in a competitive binding assay. The two protein G variants were found to interact with polyclonal IgG from different species in a similar way. Streptococcal strains expressing protein G also show affinity for human albumin, and at the molecular level protein G was found to be responsible also for the binding of albumin. Thus, both E. coli-produced protein G and the proteolytic fragment of protein G obtained from streptococci, bound albumin. On the protein G molecule, two different and separate sites were found to bind IgG and albumin. Finally, when whole streptococci were incubated with human plasma, the interactions with protein G caused a coating of the bacteria with albumin and IgG, whereas other plasma proteins showed no affinity for protein G.
authors.affiliation: Hagedorn Research Laboratory, Gentofte, Denmark.
authors.forename: L
authors.initials: L
authors.lastname: Björck
authors.forename: W
authors.initials: W
authors.lastname: Kastern
authors.forename: G
authors.initials: G
authors.lastname: Lindahl
authors.forename: K
authors.initials: K
authors.lastname: Widebäck
mesh_terms: D001426:Bacterial Proteins / Q000302:isolation & purification / Q000378:metabolism*; D001665:Binding Sites; D002846:Chromatography, Affinity; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000378:metabolism; D006801:Humans; D007074:Immunoglobulin G / Q000378:metabolism*; D012709:Serum Albumin / Q000378:metabolism*; D013291:Streptococcus / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; C034284:IgG Fc-binding protein, Streptococcus; D007074:Immunoglobulin G; D012709:Serum Albumin