Molecular Dissection of Interactions between Components of the Alternative Pathway of Complement and Decay Accelerating Factor (CD55)*

The complement regulatory protein decay accelerating factor (DAF; CD55), inhibits the alternative complement pathway by accelerating decay of the convertase enzymes formed by C3b and factor B. We show, using surface plasmon resonance, that in the absence of Mg2+, DAF binds C3b, factor B, and the Bb subunit with low affinity (KD, 14 ± 0.1, 44 ± 10, and 20 ± 7 μm, respectively). In the presence of Mg2+, DAF bound Bb or the von Willebrand factor type A subunit of Bb with higher affinities (KD, 1.3 ± 0.5 and 2.2 ± 0.1 μm, respectively). Interaction with the proenzyme C3bB was investigated by flowing factor B across a C3b-coated surface in the absence of factor D. The dissociation rate was dependent on the time of incubation, suggesting that a time-dependent conformational transition stabilized the C3b-factor B interaction. Activation by factor D (forming C3bBb) increased the complex half-life; however, the enzyme became susceptible to rapid decay by DAF, unlike the proenzyme, which was unaffected. A convertase assembled with cobra venom factor and Bb was decayed by DAF, albeit far less efficiently than C3bBb. DAF did not bind cobra venom factor, implying that Bb decay is accelerated, at least in part, through DAF binding of this subunit. It is likely that DAF binds the complex with higher affinity/avidity, promoting a conformational change in either or both subunits accelerating decay. Such analysis of component and regulator interactions will inform our understanding of inhibitory mechanisms and the ways in which regulatory proteins cooperate to control the complement cascade.

[1]  G. Smith,et al.  Biological activity, membrane‐targeting modification, and crystallization of soluble human decay accelerating factor expressed in E. coli , 2004, Protein science : a publication of the Protein Society.

[2]  O. Spiller,et al.  Mapping CD55 Function , 2003, The Journal of Biological Chemistry.

[3]  Martin Stacey,et al.  Molecular Analysis of the Epidermal Growth Factor-like Short Consensus Repeat Domain-mediated Protein-Protein Interactions , 2001, The Journal of Biological Chemistry.

[4]  M. Shoham,et al.  Molecular modeling and mechanism of action of human decay-accelerating factor. , 1996, Protein engineering.

[5]  A. Wardlaw,et al.  The properdin system and immunity. I. Demonstration and isolation of a new serum protein, properdin, and its role in immune phenomena. , 1954, Science.

[6]  A. Day,et al.  Structure-function relationships of the complement components. , 1989, Immunology today.

[7]  B. Nilsson,et al.  Complement C3b interactions studied with surface plasmon resonance technique. , 2001, International immunopharmacology.

[8]  G. Ball,et al.  Solution structure of a functionally active fragment of decay-accelerating factor , 2003, Proceedings of the National Academy of Sciences of the United States of America.

[9]  T. Fujita,et al.  Mapping of epitopes, glycosylation sites, and complement regulatory domains in human decay accelerating factor. , 1992, Journal of immunology.

[10]  D. Lublin,et al.  Efficient generation of monoclonal antibodies for specific protein domains using recombinant immunoglobulin fusion proteins: pitfalls and solutions. , 2002, Journal of immunological methods.

[11]  John D Lambris,et al.  The multifunctional role of C3: structural analysis of its interactions with physiological ligands. , 1986, Molecular immunology.

[12]  D. Stuart,et al.  Determination of the Affinity and Kinetic Constants for the Interaction between the Human Virus Echovirus 11 and Its Cellular Receptor, CD55* , 1998, The Journal of Biological Chemistry.

[13]  O. Götze,et al.  Alternative pathway of complement: recruitment of precursor properdin by the labile C3/C5 convertase and the potentiation of the pathway , 1976, The Journal of experimental medicine.

[14]  R. Levine,et al.  Interaction between the third complement protein and cell surface macromolecules. , 1977, Proceedings of the National Academy of Sciences of the United States of America.

[15]  H. Müller-Eberhard,et al.  The cobra venom factor-dependent C3 convertase of human complement. A kinetic and thermodynamic analysis of a protease acting on its natural high molecular weight substrate. , 1982, The Journal of biological chemistry.

[16]  S. Perkins,et al.  Metal-dependent conformational changes in a recombinant vWF-A domain from human factor B: a solution study by circular dichroism, fourier transform infrared and (1)H NMR spectroscopy. , 2000, Journal of Molecular Biology.

[17]  P. Nicol,et al.  The alternate pathway of complement activation. The role of C3 and its inactivator (KAF). , 1973, Immunology.

[18]  W. Dames,et al.  Complement activation by the properdin system: formation of a stoichiometric. C3 cleaving complex of properdin factor B with C36. , 1977, Immunochemistry.

[19]  P. Lachmann,et al.  The physiological breakdown of the third component of human complement. , 1980, Molecular immunology.

[20]  R. Schreiber,et al.  Formation of the initial C3 convertase of the alternative complement pathway. Acquisition of C3b-like activities by spontaneous hydrolysis of the putative thioester in native C3 , 1981, The Journal of experimental medicine.

[21]  N. Cooper FORMATION AND FUNCTION OF A COMPLEX OF THE C3 PROACTIVATOR WITH A PROTEIN FROM COBRA VENOM , 1973, The Journal of experimental medicine.

[22]  N. Cooper,et al.  The structure and function of the third component of human complement--I. The nature and extent of conformational changes accompanying C3 activation. , 1981, Molecular immunology.

[23]  Robert B Sim,et al.  Production and functional activity of a recombinant von Willebrand factor-A domain from human complement factor B. , 1999, The Biochemical journal.

[24]  D. Hourcade,et al.  Decay acceleration of the complement alternative pathway C3 convertase. , 1999, Immunopharmacology.

[25]  S. Lea Interactions of CD55 with non-complement ligands. , 2001, Biochemical Society transactions.

[26]  D. Hourcade,et al.  Decay-accelerating Factor (DAF), Complement Receptor 1 (CR1), and Factor H Dissociate the Complement AP C3 Convertase (C3bBb) via Sites on the Type A Domain of Bb* , 2002, The Journal of Biological Chemistry.

[27]  J. Lambris,et al.  Dissection of CR1, factor H, membrane cofactor protein, and factor B binding and functional sites in the third complement component. , 1996, Journal of immunology.

[28]  M. Pangburn,et al.  Nucleophilic modification of human complement protein C3: correlation of conformational changes with acquisition of C3b-like functional properties. , 1981, Biochemistry.

[29]  Z. Fishelson,et al.  Residual hemolytic and proteolytic activity expressed by Bb after decay-dissociation of C3b,Bb. , 1984, Journal of immunology.

[30]  C. Mold,et al.  Structure/function studies of human decay‐accelerating factor , 2000, Immunology.

[31]  S. Perkins,et al.  Conformational changes during the assembly of factor B from its domains by (1)H NMR spectroscopy and molecular modelling: their relevance to the regulation of factor B activity. , 2000, Journal of molecular biology.

[32]  D. Hourcade,et al.  Characterization of the Active Sites in Decay-Accelerating Factor1 , 2001, The Journal of Immunology.

[33]  Z. Fishelson,et al.  C3 convertase of the alternative complement pathway. Demonstration of an active, stable C3b, Bb (Ni) complex. , 1983, The Journal of biological chemistry.

[34]  T. Fujita,et al.  The mechanism of action of decay-accelerating factor (DAF). DAF inhibits the assembly of C3 convertases by dissociating C2a and Bb , 1987, The Journal of experimental medicine.

[35]  J. Atkinson,et al.  Competition for binding sites on C3b by CR1, CR2, MCP, factor B and factor H. , 1990, Complement and inflammation.

[36]  J. Volanakis,et al.  Surface loops adjacent to the cation-binding site of the complement factor B von Willebrand factor type A module determine C3b binding specificity. , 1997, Biochemistry.

[37]  B. Morgan,et al.  Complement Regulatory Proteins , 1999 .

[38]  Pietro Roversi,et al.  An atomic resolution model for assembly, architecture, and function of the Dr adhesins. , 2004, Molecular cell.

[39]  M. Pangburn,et al.  Differences between the binding sites of the complement regulatory proteins DAF, CR1, and factor H on C3 convertases. , 1986, Journal of immunology.

[40]  E. Pryzdial,et al.  Alternative complement pathway activation fragment Ba binds to C3b. Evidence that formation of the factor B-C3b complex involves two discrete points of contact. , 1987, The Journal of biological chemistry.

[41]  J. Atkinson,et al.  Human C3b- and C4b-regulatory proteins: a new multi-gene family. , 1985, Immunology today.

[42]  P Lukacik,et al.  Complement regulation at the molecular level: the structure of decay-accelerating factor. , 2004, Proceedings of the National Academy of Sciences of the United States of America.

[43]  S. Narayana,et al.  Complement factor D, a novel serine protease , 1996, Protein science : a publication of the Protein Society.