Publication: Recombinant generation of two fragments of the rat complement inhibitory factor H [FH(SCR1-7) and FH(SCR1-4)] and their structural and functional characterization
in comparison to FH isolated from rat serum
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Date
2006
Authors
Demberg, T. ; Heine, I. ; Götze, O. ; Altermann, W.W. ; Seliger, B. ; Schlaf, G.
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Publisher
Murcia : F. Hernández
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DOI
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info:eu-repo/semantics/article
Description
Abstract
Factor H (FH) is the predominant soluble
inhibitor of the complement system. With a
concentration of 200-800 µg/ml in human and rat plasma
it acts as a cofactor for the soluble factor I (FI)-mediated
cleavage of the component C3b to iC3b. Furthermore it
competes with factor B for binding to C3b and C3(H2O)
and promotes the dissociation of the C3bBb complex.
FH is a monomer of about 155 kDa which comprises 20
short consensus repeats (SCR), each of which is
composed of approximately 60 amino acid (aa) residues.
Two functional fragments of FH comprising the SCR1-4
or SCR1-7 were generated using either the Baculovirus
system or stably transfected human embryonal kidney
cells, respectively. These fragments, as well as FH
purified from rat serum, were first analyzed for their
relative molecular weights (Mr) using non-reducing or
reducing SDS-PAGE. The Mr of the FH variants differed
by about 20 % depending on the experimental conditions
employed. Only the Mr of proteins separated under
reducing conditions were in accordance with the MW
calculated from the aa sequence. Analyses of the
glycosylation patterns using PAS-staining showed a lack
of staining of the recombinant variants (SCR1-4 and
SCR1-7) in contrast to FH(SCR1-20) from serum. Using
a complement hemolysis assay (CH50-assay) all three
variants exhibited a molar complement inhibitory
activity of FH(1-20)/FH(1-7)/FH(1-4) of about 3/1/1. These data support the postulated model of FH bearing
three binding sites for its ligand C3b, from which one is
located in the SCR1-4, whereas the other two are located
in the SCR8-20.
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