Synergy between Two Active Sites of Human Complement Receptor Type 1 (CD35) in Complement Regulation: Implications for the Structure of the Classical Pathway C3 Convertase and Generation of More Potent Inhibitors

Krych-Goldberg, Malgorzata; Hauhart, Richard E.; Porzukowiak, Tina; Atkinson, John P.

doi:10.4049/jimmunol.175.7.4528

Cited by 22 publications

(29 citation statements)

References 54 publications

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“…Armed with this information on DAF, we then analyzed the extent of conservation of residues in DAF and in other C3 convertase regulators determined by mutagenesis to be important, either for cofactor activity or for DAA (12,18,21,22,(25)(26)(27)(28)(29)(30)(31)(32)(33)(34)(35)(36)(37)(38)(39)(40)(41). Whereas we found an absence of strict conservation of residues which distinguish the two functions, by focusing alignment on the sequence flanking the CCP2 and -3 junction where DAF function resides, we identified differences between those regulators that 1) mediate DAA and 2) mediate cofactor activity.…”

Section: Discussionmentioning

confidence: 99%

Structure-based Mapping of DAF Active Site Residues That Accelerate the Decay of C3 Convertases

Kuttner-Kondo

Hourcade

Anderson

et al. 2007

Journal of Biological Chemistry

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Focused complement activation on foreign targets depends on regulatory proteins that decay the bimolecular C3 convertases. Although this process is central to complement control, how the convertases engage and disassemble is not established. The second and third complement control protein (CCP) modules of the cell surface regulator, decay-accelerating factor (DAF, CD55), comprise the simplest structure mediating this activity. Positioning the functional effects of 31 substitution mutants of DAF CCP2 to -4 on partial structures was previously reported. In light of the high resolution crystal structure of the DAF four-CCP functional region, we now reexamine the effects of these and 40 additional mutations. Moreover, we map six monoclonal antibody epitopes and overlap their effects with those of the amino acid substitutions. The data indicate that the interaction of DAF with the convertases is mediated predominantly by two patches ϳ13 Å apart, one centered around Arg 69 and Arg 96 on CCP2 and the other around Phe 148 and Leu 171 on CCP3. These patches on the same face of the adjacent modules bracket an intermodular linker of critical length (16 Å ). Although the key DAF residues in these patches are present or there are conservative substitutions in all other C3 convertase regulators that mediate decay acceleration and/or provide factor I-cofactor activity, the linker region is highly conserved only in the former. Intra-CCP regions also differ. Linker region comparisons suggest that the active CCPs of the decay accelerators are extended, whereas those of the cofactors are tilted. Intra-CCP comparisons suggest that the two classes of regulators bind different regions on their respective ligands.The C3 convertases of the classical and alternative pathways are the central enzymes of the complement cascade (1). These bimolecular complexes, C4b2a and C3bBb, produce anaphylatoxin C3a, locally amplify C3b deposition, and serve as sites for the assembly of the C5 convertases, C4b2a3b and C3bBb3b. These trimeric convertases, in turn, generate anaphylatoxin C5a and initiate the terminal pathway, leading to formation of lytic C5b-9 membrane attack complexes. The relative rates of assembly and disassembly of the C3 convertases lie at the heart of complement regulation, and their physiologic modulation ensures that complement acts in a proportionate and targeted fashion.To focus complement activation on foreign targets, prevent nearby activation in the fluid phase, and simultaneously protect self tissues from complement-mediated injury, the C3 convertases are controlled by both serum-and cell-associated regulatory proteins (2, 3). Because the convertases assemble on C4b and C3b fragments that condense indiscriminately with free hydroxyl and amino groups on foreign targets and these same acceptor groups are present on all biological membranes, their formation on self cells at the single enzyme level cannot be avoided. To prevent further propagation of the cascade, which would induce self cell injury, self cells possess decay-acc...

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Section: Discussionmentioning

confidence: 99%

Structure-based Mapping of DAF Active Site Residues That Accelerate the Decay of C3 Convertases

Kuttner-Kondo

Hourcade

Anderson

et al. 2007

Journal of Biological Chemistry

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“…Our identification of a short peptide within this N-terminal domain may be helpful for future design of smaller peptidomimetic molecules that can replace the larger 3 SCRs in APT070. Interestingly, mutations that considerably increase decay acceleration for the classical pathway C3 convertase have been identified that map to residues within our peptide 4 [20]. By making synthetic peptides harboring such mutations we plan on testing their effectiveness in preventing complement-mediated RBC destruction using our assays.…”

Section: Discussionmentioning

confidence: 99%

“…Previous mutagenesis data of SCRs-1-4 have identified several critical residues for decay accelerating activities in the N-terminal domain [16][17][18][19][20]. For example, three positively charged amino acids Arg59, Arg60, and Lys61 at SCR-1/SCR-2 junction, Arg64, Asn65, Thr103, and Thr110 in SCR-2 as well as Gly35 in SCR-1 were found to be important for both decay accelerating activity while Phe82 was required primarily for decay accelerating activity [20]. To date, no structural data for the N-terminal domain of CR1 is available.…”

Section: Author Manuscriptmentioning

confidence: 99%

Identification of a complement receptor 1 peptide for inhibition of immune hemolysis

Yu¹,

Heck²,

Debnath³

et al. 2007

Biochemical and Biophysical Research Communications

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Complement sensitization of red blood cells (RBCs) can cause life-threatening hemolytic anemias. We have previously shown that complement receptor 1 (CR1) derivatives specifically the Nterminal region with decay accelerating activity (DAA) for inactivation of a key enzyme in the complement cascade can reduce complement-mediated RBC destruction in vitro and in an in vivo mouse model of hemolytic transfusion reaction. In the present study, we have modeled the Nterminal CR1 molecule based on the X-ray crystal structure of decay accelerating factor and the NMR structure of a homologous CR1 domain. Based on the homology model, we identified a 34-mer peptide encompassing the putative DAA which in vitro reduced hemolysis, C3a release and surface C3 deposition. More importantly, this peptide at 0.6 mM was effective in prolonging survival of transfused incompatible RBCs in vivo. Our results indicate that CR1-based structurefunction studies may provide insights for developing structure-derived transfusion therapeutics in the future. KeywordsComplement receptor 1; CD35; Complement inhibition; Transfusion therapy; Hemolysis; Homology modeling; Mouse model; Transfusion reactions; CR1 peptides; C3a; C3bThe complement system is an important part of the innate immune system for fighting infections and foreign molecules before an adaptive response is developed. To execute its functions, the system has to be activated. A key step in the complement activation cascade is the formation of the C3 convertase complex on the surface of target cells which cleaves C3 into C3b, the main effector of complement, and C3a, a potent anaphylatoxin that is released into the medium. C3b and its degradation products on target cells act as opsonins marking them for removal from the circulation by phagocytes. In addition, if sufficient quantities of ☆ This study was supported in part by a grants (to K.Y.) from the NIH R01 HL69102, and the American Heart Association Grant-inAid Heritage Affiliate. * Corresponding author. Fax: +1 212 570 3195. kyazdanbakhsh@nybloodcenter.org (K. Yazdanbakhsh).. HHS Public Access Author Manuscript Author ManuscriptAuthor Manuscript Author ManuscriptC3b are formed on the cell surface, they can result in serial activation of complement proteins (C5-C9) that form the C5b-9 complex, also known as the membrane attack complex. Once adequate number of C5b-9 complexes form, the target cell is lysed. As expected, inappropriate complement activation can have deleterious effects. In the transfusion medicine setting, complement sensitization of red blood cells (RBCs) can result in life-threatening hemolytic transfusion reactions as well as hemolytic anemias [1]. There is thus a critical need for a therapeutically applicable complement inhibitor. To date several inhibitors have been described [2], but none of these has yet been adopted as a therapeutic agent. A strategy to develop complement inhibitors has been to manipulate naturally occurring complement regulatory proteins that act at various steps in the activation casca...

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“…Thus, flexibility is likely important for activity in some RCAs, and in particular for the larger ones with multiple binding sites such as factor H and CR1 (and indeed CR2). In this respect, it is probably no coincidence that engineering of RCAs with residues inserted into linkers normally devastates function, that linker residues and intermodular "joints" of RCAs frequently contribute to intermolecular interactions, and that binding sites are often composite involving sub-sites on adjacent modules (Blom et al, 2000;Brodbeck et al, 2000;Jenkins et al, 2005;Krych-Goldberg and Atkinson, 2001;Krych-Goldberg et al, 2005). By binding at the intermodular linker or across multiple modules, ligands will bridge adjacent modules and modulate the overall structure of an RCA.…”

Section: Structure-function Relationships In the Regulators Of Complementioning

confidence: 99%

Deciphering complement mechanisms: The contributions of structural biology

Arlaud

Barlow

Gaboriaud

et al. 2007

Molecular Immunology

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Since the resolution of the first three-dimensional structure of a complement component in 1980, considerable efforts have been put into the investigation of this system through structural biology techniques, resulting in about a hundred structures deposited in the Protein Data Bank by the beginning of 2007. By revealing its mechanisms at the atomic level, these approaches significantly improve our understanding of complement, opening the way to the rational design of specific inhibitors. This review is co-authored by some of the researchers currently involved in the structural biology of complement and its purpose is to illustrate, through representative examples, how X-ray crystallography and NMR techniques help us decipher the many sophisticated mechanisms that underlie complement functions.

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Synergy between Two Active Sites of Human Complement Receptor Type 1 (CD35) in Complement Regulation: Implications for the Structure of the Classical Pathway C3 Convertase and Generation of More Potent Inhibitors

Cited by 22 publications

References 54 publications

Structure-based Mapping of DAF Active Site Residues That Accelerate the Decay of C3 Convertases

Structure-based Mapping of DAF Active Site Residues That Accelerate the Decay of C3 Convertases

Identification of a complement receptor 1 peptide for inhibition of immune hemolysis

Deciphering complement mechanisms: The contributions of structural biology

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