Single-step purification of human C4b-binding protein (C4BP) by affinity chromatography on a peptide derived from a streptococcal surface protein

• Published in: Journal of immunological methods Vol. 297; no. 1; pp. 83 - 95
• Main Authors: Persson, Jenny, Lindahl, Gunnar
• Format: Journal Article
• Language: English
• Published: Amsterdam Elsevier B.V 01.02.2005; Elsevier
• Subjects: Amino Acid Sequence; Animals; Antigens, Bacterial - chemistry; Bacterial Outer Membrane Proteins - chemistry; Bacteriological methods and techniques used in bacteriology; Bacteriology; Basic Medicine; Biological and medical sciences; C4BP; Carrier Proteins - chemistry; Cattle; Chromatography, Affinity - methods; Complement C4b-Binding Protein; Depletion; Dimerization; Fundamental and applied biological sciences. Psychology; Fundamental immunology; Histocompatibility Antigens - blood; Histocompatibility Antigens - chemistry; Histocompatibility Antigens - isolation & purification; Humans; Immunologi inom det medicinska området (Här ingår: Cell- och immunterapi); Immunology in the Medical Area (including Cell and Immunotherapy); Medical and Health Sciences; Medicin och hälsovetenskap; Medicinska och farmaceutiska grundvetenskaper; Mice; Microbiology; Molecular immunology; Molecular Sequence Data; Peptide; Peptides - chemistry; Protein purification; Rabbits; Streptococcal M protein; Streptococcus; Techniques; C4BP; CCP; CD; SCR; Depletion; Protein purification; Streptococcal M protein; Peptide; Human; Purification; Peptides; Immunological method; Infection; Streptococcaceae; Binding protein; Affinity chromatography; Streptococcal infection; Streptococcus A; Bacteriosis; Bacteria; Micrococcales
• ISSN: 0022-1759; 1872-7905; 1872-7905
• DOI: 10.1016/j.jim.2004.11.024

Many Gram-positive bacteria express surface proteins that bind human plasma proteins. These bacterial proteins, and derivatives of them, are of interest for analysis of bacterial pathogenesis and as immunochemical tools. Well-characterized examples include the IgG-binding reagents staphylococcal protein A and streptococcal protein G, and the recently described streptococcal IgA-binding peptide Sap. Here, we show that a peptide derived from the streptococcal M22 protein can be used for single-step affinity purification of the human complement regulator C4b-binding protein (C4BP). Binding of C4BP was strongly enhanced by dimerization of the peptide via a C-terminal cysteine residue not present in the intact M22 protein. The purified C4BP had the expected binding characteristics, and acted as a cofactor for factor I in the degradation of C4b. Passage of serum through a peptide column under non-saturating conditions resulted in binding of >99.5% of serum C4BP, implying that such a column can be used to deplete serum of C4BP. These data indicate that the C4BP-binding peptide is a versatile tool that can be used for simple and rapid purification of biologically active human C4BP or for removal of C4BP from serum.