Isolation and characterization of a 14-kDa albumin-binding fragment of streptococcal protein G.
Protein G, a streptococcal cell wall protein, has separate binding sites for human albumin and IgG. Streptococci expressing protein G were treated with the bacteriolytic agent mutanolysin. Several IgG- and human serum albumin (HSA)-binding peptides were identified in the material thus solubilized and one of these, a 14-kDa peptide, was found to bind HSA but not IgG in Western blot experiments. This molecule was purified by affinity chromatography on Sepharose coupled with HSA followed by gel filtration on Sepharose 6B and a final affinity chromatography on IgG-Sepharose, by which low Mr W(15 to 20 kDa)IgG-binding peptides were removed. In different binding experiments the purified 14-kDa peptide bound exclusively HSA and the equilibrium constant between the peptide and HSA was determined to be 3.4 X 10(8) M-1. The relation between the 14-kDa molecule and protein G was studied by analyzing the N-terminal amino acid sequence of the peptide and comparing it with the previously determined protein G sequence. The 40 N-terminal amino acids were found to be identical with an amino acid sequence starting at position 62 in the protein G molecule. These and previous data enabled us to locate the albumin binding to the repetitively arranged domains in the N-terminal half of the protein G molecule.
Sjöbring
U
U
Department of Medical Microbiology, University of Lund, Sweden.
Falkenberg
C
C
Nielsen
E
E
Akerström
B
B
Björck
L
L
D001426:Bacterial Proteins / Q000302:isolation & purification*; D002352:Carrier Proteins / Q000302:isolation & purification*; D006801:Humans; D007074:Immunoglobulin G / Q000378:metabolism; D008970:Molecular Weight; D010446:Peptide Fragments / Q000302:isolation & purification*; D017457:Receptors, Albumin; D011956:Receptors, Cell Surface / Q000032:analysis*; D012709:Serum Albumin / Q000276:immunology / Q000378:metabolism*; D013291:Streptococcus / Q000032:analysis* / Q000276:immunology / Q000378:metabolism
D001426:Bacterial Proteins; D002352:Carrier Proteins; C034284:IgG Fc-binding protein, Streptococcus; D007074:Immunoglobulin G; D010446:Peptide Fragments; D017457:Receptors, Albumin; D011956:Receptors, Cell Surface; D012709:Serum Albumin