Abstract
Human C8 is one of five components of the membrane attack complex of complement (MAC). It is composed of a disulfide-linked C8alpha-gamma heterodimer and a noncovalently associated C8beta chain. The C8alpha and C8beta subunits contain a pair of N-terminal modules [thrombospondin type 1 (TSP1) + low-density lipoprotein receptor class A (LDLRA)] and a pair of C-terminal modules [epidermal growth factor (EGF) + TSP1]. The middle segment of each protein is referred to as the membrane attack complex/perforin domain (MACPF). During MAC formation, C8alpha mediates binding and self-polymerization of C9 to form a pore-like structure on the membrane of target cells. In this study, the portion of C8alpha involved in binding C9 was identified using recombinant C8alpha constructs in which the N- and/or C-terminal modules were either exchanged with those from C8beta or deleted. Those constructs containing the C8alpha N-terminal TSP1 or LDLRA module together with the C8alpha MACPF domain retained the ability to bind C9 and express C8 hemolytic activity. By contrast, those containing the C8alpha MACPF domain alone or the C8alpha MACPF domain and C8alpha C-terminal modules lost this ability. These results indicate that both N-terminal modules in C8alpha have a role in forming the principal binding site for C9 and that binding may be dependent on a cooperative interaction between these modules and the C8alpha MACPF domain.
Publication types
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Research Support, U.S. Gov't, P.H.S.
MeSH terms
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Animals
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Binding Sites / genetics
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COS Cells
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Complement C8 / chemistry*
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Complement C8 / genetics
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Complement C8 / metabolism
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Complement C8 / physiology
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Complement C9 / chemistry*
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Complement C9 / metabolism
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Complement Membrane Attack Complex / chemistry
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Complement Membrane Attack Complex / genetics
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Complement Membrane Attack Complex / metabolism
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Genetic Vectors / chemical synthesis
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Genetic Vectors / metabolism
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Genetic Vectors / physiology
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Humans
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Membrane Glycoproteins / chemistry
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Membrane Glycoproteins / genetics
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Membrane Glycoproteins / metabolism
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Peptide Fragments / chemistry*
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Peptide Fragments / genetics
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Peptide Fragments / metabolism
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Peptide Fragments / physiology
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Perforin
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Pore Forming Cytotoxic Proteins
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Protein Binding / genetics
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Protein Structure, Tertiary / genetics
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Protein Subunits / chemistry*
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Protein Subunits / genetics
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Protein Subunits / metabolism
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Receptors, LDL / classification
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Receptors, LDL / genetics
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Receptors, LDL / metabolism
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Recombinant Fusion Proteins / chemistry
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Recombinant Fusion Proteins / metabolism
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Sequence Deletion / genetics
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Thrombospondin 1 / chemistry
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Thrombospondin 1 / genetics
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Thrombospondin 1 / metabolism
Substances
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Complement C8
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Complement C9
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Complement Membrane Attack Complex
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Membrane Glycoproteins
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Peptide Fragments
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Pore Forming Cytotoxic Proteins
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Protein Subunits
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Receptors, LDL
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Recombinant Fusion Proteins
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Thrombospondin 1
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Perforin