Biomedical subjects

Jens Hellwage

Publications and source records attributed to Jens Hellwage.

5 recordsLinked to original sources

PubMed2003

Immune evasion of Borrelia burgdorferi: mapping of a complement-inhibitor factor H-binding site of BbCRASP-3, a novel member of the Erp protein family.

The causative agents of Lyme disease, Borrelia burgdorferi s.s., B. garinii, and B. afzelii, differ in their susceptibility to complement-mediated lysis. This phenomenon apparently depends on the expression of proteins termed complement regulator-acquiring surface proteins (CRASP) and their binding to the inhibitory plasma proteins factor H and FHL-1. To characterize these bacterial proteins in more detail we have now isolated from a B. burgdorferi expression library a novel factor H-binding protein. In accordance with our previous studies this protein was termed BbCRASP-3 and represents a novel member of the polymorphic Erp (OspE/F-related) protein family. On the basis of protease accessibility assays using intact spirochetes, BbCRASP-3 is identified as a surface-exposed protein and binds the C-terminal short consensus repeats of factor H. Applying deletion mutants of BbCRASP-3, the factor H-binding site was mapped to the nine-amino-acid motif LEVLKKNLK localized at the C-terminal end of BbCRASP-3. Factor H bound to BbCRASP-3 maintains its cofactor activity in factor I-mediated C3b inactivation. Binding of BbCRASP-3 to factor H can be inhibited by heparin, a physiological ligand of the complement regulator factor H. Blocking of factor-H-binding by soluble BbCRASP-3 leads to an increase of complement deposition on intermediate serum-resistant strain ZS7. In conclusion, BbCRASP-3 has been identified as a novel factor H-binding protein on B. burgdorferi which by conferring complement resistance to the pathogen may contribute to its persistence in the mammalian host.

Amino Acid Sequence↗

PubMed2003

Mutations in factor H reduce binding affinity to C3b and heparin and surface attachment to endothelial cells in hemolytic uremic syndrome.

Hemolytic uremic syndrome (HUS) is a disease characterized by microangiopathic hemolytic anemia, thrombocytopenia, and acute renal failure. Recent studies have identified a factor H-associated form of HUS, caused by gene mutations that cluster in the C-terminal region of the complement regulator factor H. Here we report how three mutations (E1172Stop, R1210C, and R1215G; each of the latter two identified in three independent cases from different, unrelated families) affect protein function. All three mutations cause reduced binding to the central complement component C3b/C3d to heparin, as well as to endothelial cells. These defective features of the mutant factor H proteins explain progression of endothelial cell and microvascular damage in factor H-associated genetic HUS and indicate a protective role of factor H for tissue integrity during thrombus formation.

Animals↗

PubMed2002

Complement C3b/C3d and cell surface polyanions are recognized by overlapping binding sites on the most carboxyl-terminal domain of complement factor H.

Factor H (FH) is a potent suppressor of the alternative pathway of C in plasma and when bound to sialic acid- or glycosaminoglycan-rich surfaces. Of the three interaction sites on FH for C3b, one interacts with the C3d part of C3b. In this study, we generated recombinant constructs of FH and FH-related proteins (FHR) to define the sites required for binding to C3d. In FH, the C3d-binding site was localized by surface plasmon resonance analysis to the most C-terminal short consensus repeat domain (SCR) 20. To identify amino acids of FH involved in binding to C3d and heparin, we compared the sequences of FH and FHRs and constructed a homology-based molecular model of SCR19-20 of FH. Subsequently, we created an SCR15-20 mutant with substitutions in five amino acids that were predicted to be involved in the binding interactions. These mutations reduced binding of the SCR15-20 construct to both C3b/C3d and heparin. Binding of the wild-type SCR15-20, but not the residual binding of the mutated SCR15-20, to C3d was inhibited by heparin. This indicates that the heparin- and C3d-binding sites are overlapping. Our results suggest that a region in the most C-terminal domain of FH is involved in target recognition by binding to C3b and surface polyanions. Mutations in this region, as recently reported in patients with familial hemolytic uremic syndrome, may lead to indiscriminatory C attack against self cells.

Amino Acid Sequence↗

PubMed2002

Onchocerca volvulus microfilariae avoid complement attack by direct binding of factor H.

The filarial parasite Onchocerca volvulus is the causative agent of river blindness. The adult worms produce microfilariae (mf), which are responsible for the disease pathogenesis; mf activate the complement system, but the activation stops before the formation of terminal complement complexes. Because of the arrest of complement activation, this study analyzed binding of the main alternative pathway regulator, factor H (fH), to the mf. The mf bound fH after incubation in nonimmune human serum or with purified radiolabeled fH. In the presence of factor I, mf-bound fH promoted the cleavage of complement 3 molecule b (C3b) to iC3b. An analysis with recombinant constructs of fH showed that the C-terminal short consensus repeats (SCRs) 8-20 of fH bound to mf, whereas the N-terminal SCRs 1-7 containing the complement-regulatory domains in SCRs 1-5 did not. Thus, mf of the nematode O. volvulus may evade human complement by binding fH and by promoting inactivation of C3b into iC3b.

Animals↗

PubMed2002

Streptococcus pneumoniae evades complement attack and opsonophagocytosis by expressing the pspC locus-encoded Hic protein that binds to short consensus repeats 8-11 of factor H.

Streptococcus pneumoniae is an important cause of upper and lower respiratory tract infections, meningitis, peritonitis, bacterial arthritis, and sepsis. Here we have studied a novel immune evasion mechanism of serotype 3 pneumococci, which are particularly resistant to phagocytosis. On their surfaces the bacteria express the factor H-binding inhibitor of complement (Hic), a protein of the pneumococcal surface protein C family. Using radioligand binding, microtiter plate assays, surface plasmon resonance analysis, and recombinant constructs of factor H, we located the binding site of Hic to short consensus repeats (SCRs) 8-11 in the middle part of factor H. This represents a novel microbial interaction region on factor H. The only other ligand known so far for SCRs 8-11 of factor H is C-reactive protein (CRP), an acute phase protein that binds to the pneumococcal C-polysaccharide. The binding sites of Hic and CRP within the SCR8-11 region were different, however, because CRP did not inhibit the binding of Hic and required calcium for binding. Binding of factor H to Hic-expressing pneumococci promoted factor I-mediated cleavage of C3b and restricted phagocytosis of pneumococci. Thus, virulent pneumococci avoid complement attack and opsonophagocytosis by recruiting functionally active factor H with the Hic surface protein. Hic binds to a previously unrecognized microbial interaction site in the middle part of factor H.

Bacterial Proteins↗