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Involvement of amino acid residues 423-429 of human protein S in binding to C4b-binding protein. 1998

J A Fernández, and J H Griffin, and G T Chang, and J Stam, and P H Reitsma, and R M Bertina, and B N Bouma
Department of Molecular and Experimental Medicine, Scripps Research Institute, La Jolla, California 92037, USA.

Human protein S binds to C4b-binding protein (C4BP) both in plasma and in a system using purified proteins. Amino acid residues 420-434 of the first disulfide loop of the sex hormone binding globulinlike domain of protein S are involved in the interaction of protein S with C4BP. To define the involvement of specific polar amino acids within residues 420-434, we studied in parallel synthetic protein S peptides and recombinant protein S variants containing the same amino acid replacements, K423E, E424K, Q427E and K429E. Synthetic peptide analogs of peptide PSP-420 (residues 420-434) were assayed for binding C4BP and as inhibitors of complex formation. The PSP-420 peptide and the analogous peptide with the substitution E424K, but not the peptides containing the substitutions K423E and K429E, were able to bind C4BP. Recombinant proteins with mutations of K423E, Q427E and K429E showed reduced affinity for C4BP compared to plasma protein S, recombinant wild type protein S, or E424K-protein S. These results suggest that Lys-423, Gln-427 and Lys-429 of protein S are important for normal binding to C4BP. The anti-protein S monoclonal antibody LJ-56, raised against peptide PSP-420, recognizes only free protein S and inhibits complex formation with C4BP. Antibody LJ-56 recognized the E424K and Q427E peptides but not the K423E or K429E peptides. Similarly, the E424K and Q427E protein S mutants were recognized by LJ-56, whereas the K423E and K429E protein S mutants were not recognized. This suggests that both in the peptide PSP-420 and in protein S, Lys-423 and Lys-429 significantly contribute to binding to antibody LJ-56. These results demonstrate that protein S residues 423, 427 and 429, but not residue 424, are involved in binding to both the antibody LJ-56 and to C4BP. When peptides PSP 420 and SL-6 (residues 447-460) with carboxyterminal amide or carboxylate moieties were compared to their ability to inhibit C4BP-protein S complexation, PSP-420-amide was the most potent. This finding together with the other results described here supports the hypothesis that the residues 420 and 434 in protein S provides a major binding site for C4BP.

UI MeSH Term Description Entries
D008969 Molecular Sequence Data Descriptions of specific amino acid, carbohydrate, or nucleotide sequences which have appeared in the published literature and/or are deposited in and maintained by databanks such as GENBANK, European Molecular Biology Laboratory (EMBL), National Biomedical Research Foundation (NBRF), or other sequence repositories. Sequence Data, Molecular,Molecular Sequencing Data,Data, Molecular Sequence,Data, Molecular Sequencing,Sequencing Data, Molecular
D010446 Peptide Fragments Partial proteins formed by partial hydrolysis of complete proteins or generated through PROTEIN ENGINEERING techniques. Peptide Fragment,Fragment, Peptide,Fragments, Peptide
D011485 Protein Binding The process in which substances, either endogenous or exogenous, bind to proteins, peptides, enzymes, protein precursors, or allied compounds. Specific protein-binding measures are often used as assays in diagnostic assessments. Plasma Protein Binding Capacity,Binding, Protein
D011951 Receptors, Complement Molecules on the surface of some B-lymphocytes and macrophages, that recognize and combine with the C3b, C3d, C1q, and C4b components of complement. Complement Receptors,Complement Receptor,Complement Receptor Type 1,Receptor, Complement
D011993 Recombinant Fusion Proteins Recombinant proteins produced by the GENETIC TRANSLATION of fused genes formed by the combination of NUCLEIC ACID REGULATORY SEQUENCES of one or more genes with the protein coding sequences of one or more genes. Fusion Proteins, Recombinant,Recombinant Chimeric Protein,Recombinant Fusion Protein,Recombinant Hybrid Protein,Chimeric Proteins, Recombinant,Hybrid Proteins, Recombinant,Recombinant Chimeric Proteins,Recombinant Hybrid Proteins,Chimeric Protein, Recombinant,Fusion Protein, Recombinant,Hybrid Protein, Recombinant,Protein, Recombinant Chimeric,Protein, Recombinant Fusion,Protein, Recombinant Hybrid,Proteins, Recombinant Chimeric,Proteins, Recombinant Fusion,Proteins, Recombinant Hybrid
D003169 Complement Inactivator Proteins Serum proteins that negatively regulate the cascade process of COMPLEMENT ACTIVATION. Uncontrolled complement activation and resulting cell lysis is potentially dangerous for the host. The complement system is tightly regulated by inactivators that accelerate the decay of intermediates and certain cell surface receptors. Complement Cytolysis Inhibiting Proteins,Complement Cytolysis Inhibitor Proteins,Complement Inactivating Proteins,Serum Complement Inactivators,Complement Inactivators, Serum,Inactivating Proteins, Complement,Inactivator Proteins, Complement,Inactivators, Serum Complement,Proteins, Complement Inactivating,Proteins, Complement Inactivator
D006023 Glycoproteins Conjugated protein-carbohydrate compounds including MUCINS; mucoid, and AMYLOID glycoproteins. C-Glycosylated Proteins,Glycosylated Protein,Glycosylated Proteins,N-Glycosylated Proteins,O-Glycosylated Proteins,Glycoprotein,Neoglycoproteins,Protein, Glycosylated,Proteins, C-Glycosylated,Proteins, Glycosylated,Proteins, N-Glycosylated,Proteins, O-Glycosylated
D006801 Humans Members of the species Homo sapiens. Homo sapiens,Man (Taxonomy),Human,Man, Modern,Modern Man
D000595 Amino Acid Sequence The order of amino acids as they occur in a polypeptide chain. This is referred to as the primary structure of proteins. It is of fundamental importance in determining PROTEIN CONFORMATION. Protein Structure, Primary,Amino Acid Sequences,Sequence, Amino Acid,Sequences, Amino Acid,Primary Protein Structure,Primary Protein Structures,Protein Structures, Primary,Structure, Primary Protein,Structures, Primary Protein
D000911 Antibodies, Monoclonal Antibodies produced by a single clone of cells. Monoclonal Antibodies,Monoclonal Antibody,Antibody, Monoclonal

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