Staphylococcus aureus SdrE captures complement factor H's C-terminus via a novel ‘close, dock, lock and latch' mechanism for complement evasion

Complement factor H (CFH) is a soluble complement regulatory protein essential for the down-regulation of the alternative pathway on interaction with specific markers on the host cell surface. It recognizes the complement component 3b (C3b) and 3d (C3d) fragments in addition to self cell markers (i.e. glycosaminoglycans, sialic acid) to distinguish host cells that deserve protection from pathogens that should be eliminated. The Staphylococcus aureus surface protein serine–aspartate repeat protein E (SdrE) was previously reported to bind human CFH as an immune-evasion tactic. However, the molecular mechanism underlying SdrE–CFH-mediated immune evasion remains unknown. In the present study, we identified a novel region at CFH's C-terminus (CFH1206–1226), which binds SdrE N2 and N3 domains (SdrEN2N3) with high affinity, and determined the crystal structures of apo-SdrEN2N3 and the SdrEN2N3–CFH1206–1226 complex. Comparison of the structure of the CFH–SdrE complex with other CFH structures reveals that CFH's C-terminal tail flips from the main body to insert into the ligand-binding groove of SdrE. In addition, SdrEN2N3 adopts a ‘close’ state in the absence of CFH, which undergoes a large conformational change on CFH binding, suggesting a novel ‘close, dock, lock and latch' (CDLL) mechanism for SdrE to recognize its ligand. Our findings imply that SdrE functions as a ‘clamp' to capture CFH's C-terminal tail via a unique CDLL mechanism and sequesters CFH on the surface of S. aureus for complement evasion.

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Cross-Reactivity of Antipneumococcal Surface Protein C (PspC) Antibodies with Different Strains and Evaluation of Inhibition of Human Complement Factor H and Secretory IgA Binding via PspC

Clinical and Vaccine Immunology ◽

10.1128/cvi.05706-11 ◽

2012 ◽

Vol 19 (4) ◽

pp. 499-507 ◽

Cited By ~ 11

Author(s):

Adriana T. Moreno ◽

Maria Leonor S. Oliveira ◽

Paulo L. Ho ◽

Cintia F. M. Vadesilho ◽

Giovana M. P. Palma ◽

...

Keyword(s):

Flow Cytometry ◽

Western Blotting ◽

Protein C ◽

Alternative Pathway ◽

Surface Protein ◽

Cross Reactivity ◽

Factor H ◽

Secretory Iga ◽

Complement Factor ◽

Cell Extracts

ABSTRACTPneumococcal surface protein C (PspC) is an important candidate for a cost-effective vaccine with broad coverage against pneumococcal diseases. Previous studies have shown thatStreptococcus pneumoniaeis able to bind to both human factor H (FH), an inhibitor of complement alternative pathway, and human secretory IgA (sIgA) via PspC. PspC was classified into 11 groups based on variations of the gene. In this work, we used three PspC fragments from different groups (PspC3, PspC5, and PspC8) to immunize mice for the production of antibodies. Immunization with PspC3 induced antibodies that recognized the majority of the clinical isolates as analyzed by Western blotting of whole-cell extracts and flow cytometry of intact bacteria, while anti-PspC5 antibodies showed cross-reactivity with the paralogue pneumococcal surface protein A (PspA), and anti-PspC8 antibodies reacted only with the PspC8-expressing strain. Most of the isolates tested showed strong binding to FH and weaker interaction with sIgA. Preincubation with anti-PspC3 and anti-PspC5 IgG led to some inhibition of binding of FH, and preincubation with anti-PspC3 partially inhibited sIgA binding in Western blotting. The analysis of intact bacteria through flow cytometry showed only a small decrease in FH binding after incubation of strain D39 with anti-PspC3 IgG, and one clinical isolate showed inhibition of sIgA binding by anti-PspC3 IgG. We conclude that although anti-PspC3 antibodies were able to recognize PspC variants from the majority of the strains tested, partial inhibition of FH and sIgA binding through anti-PspC3 antibodiesin vitrocould be observed for only a restricted number of isolates.

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Cleavage of Complement C3b to iC3b on the Surface of Staphylococcus aureus Is Mediated by Serum Complement Factor I

Infection and Immunity ◽

10.1128/iai.72.5.2858-2863.2004 ◽

2004 ◽

Vol 72 (5) ◽

pp. 2858-2863 ◽

Cited By ~ 36

Author(s):

K. M. Cunnion ◽

P. S. Hair ◽

E. S. Buescher

Keyword(s):

Staphylococcus Aureus ◽

Human Serum ◽

Serum Proteins ◽

Regulatory Protein ◽

Factor H ◽

Complement Factor ◽

Serum Factor ◽

Complement Protein ◽

Factor I ◽

Serotype 5

ABSTRACT Complement-mediated opsonization of Staphylococcus aureus bearing the dominant capsule serotypes, serotypes 5 and 8, remains incompletely understood. We have previously shown that complement plays a vital role in the efficient phagocytosis of a serotype 5 S. aureus strain and that the opsonic fragments of the central complement protein C3, C3b and iC3b, were present on the bacterial surface after incubation in human serum. In the present studies, C3b and iC3b were found on several serotype 5 and 8 S. aureus strains after incubation in human serum. Using purified classical activation pathway complement proteins and the Western blot assay, we showed that when C3b was generated on the S. aureus surface no iC3b fragments were found, suggesting that other serum proteins may be required for cleaving C3b to iC3b. When C3b-coated S. aureus was incubated with serum factor I, a complement regulatory protein, iC3b was generated. Purified factor H, a serum protein cofactor for factor I, did not enhance factor I-mediated cleavage of C3b. These findings suggest that C3b cleavage to iC3b on S. aureus is mediated by serum factor I and does not require factor H.

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Clumping Factor A Interaction with Complement Factor I Increases C3b Cleavage on the Bacterial Surface of Staphylococcus aureus and Decreases Complement-Mediated Phagocytosis

Infection and Immunity ◽

10.1128/iai.01065-09 ◽

2010 ◽

Vol 78 (4) ◽

pp. 1717-1727 ◽

Cited By ~ 65

Author(s):

Pamela S. Hair ◽

Charlene G. Echague ◽

Amber M. Sholl ◽

Justin A. Watkins ◽

Joan A. Geoghegan ◽

...

Keyword(s):

Staphylococcus Aureus ◽

Surface Protein ◽

Factor H ◽

Complement Factor ◽

Wild Type ◽

Bacterial Surface ◽

Complement Control Protein ◽

Factor I ◽

Protein Factor ◽

Fibrinogen Binding

ABSTRACT The human complement system is important in the immunological control of Staphylococcus aureus infection. We showed previously that S. aureus surface protein clumping factor A (ClfA), when expressed in recombinant form, bound complement control protein factor I and increased factor I cleavage of C3b to iC3b. In the present study, we show that, compared to the results for the wild type, when isogenic ClfA-deficient S. aureus mutants were incubated in serum, they bound less factor I, generated less iC3b on the bacterial surface, and bound fewer C3 fragments. It has been shown previously that two amino acids in ClfA (P336 and Y338) are essential for fibrinogen binding. However, S. aureus expressing ClfA(P336A Y338S) was less virulent than ClfA-deficient strains in animal models. This suggested that ClfA contributed to S. aureus virulence by a mechanism different than fibrinogen binding. In the present study, we showed that S. aureus expressing ClfA(P336A Y338S) was more susceptible to complement-mediated phagocytosis than a ClfA-null mutant or the wild type. Unlike ClfA, ClfA(P336A Y338S) did not enhance factor I cleavage of C3b to iC3b and inhibited the cofactor function of factor H. Fibrinogen enhanced factor I binding to ClfA and the S. aureus surface. Twenty clinical S. aureus strains all expressed ClfA and bound factor I. High levels of factor I binding by clinical strains correlated with poor phagocytosis. In summary, our results suggest that the interaction of ClfA with factor I contributes to S. aureus virulence by a complement-mediated mechanism.

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Borrelia burgdorferi Regulates Expression of Complement Regulator-Acquiring Surface Protein 1 during the Mammal-Tick Infection Cycle

Infection and Immunity ◽

10.1128/iai.73.11.7398-7405.2005 ◽

2005 ◽

Vol 73 (11) ◽

pp. 7398-7405 ◽

Cited By ~ 56

Author(s):

Kate von Lackum ◽

Jennifer C. Miller ◽

Tomasz Bykowski ◽

Sean P. Riley ◽

Michael E. Woodman ◽

...

Keyword(s):

Borrelia Burgdorferi ◽

Immune Evasion ◽

Expression Patterns ◽

Surface Protein ◽

Factor H ◽

Complement Factor ◽

Human Pathogens ◽

Infection Cycle ◽

Tick Infection ◽

Complement Regulator

ABSTRACT During the natural mammal-tick infection cycle, the Lyme disease spirochete Borrelia burgdorferi comes into contact with components of the alternative complement pathway. B. burgdorferi, like many other human pathogens, has evolved the immune evasion strategy of binding two host-derived fluid-phase regulators of complement, factor H and factor H-like protein 1 (FHL-1). The borrelial complement regulator-acquiring surface protein 1 (CRASP-1) is a surface-exposed lipoprotein that binds both factor H and FHL-1. Analysis of CRASP-1 expression during the mammal-tick infectious cycle indicated that B. burgdorferi expresses this protein during mammalian infection, supporting the hypothesized role for CRASP-1 in immune evasion. However, CRASP-1 synthesis was repressed in bacteria during colonization of vector ticks. Analysis of cultured bacteria indicated that CRASP-1 is differentially expressed in response to changes in pH. Comparisons of CRASP-1 expression patterns with those of other infection-associated B. burgdorferi proteins, including the OspC, OspA, and Erp proteins, indicated that each protein is regulated through a unique mechanism.

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Hiding in plain sight: immune evasion by the staphylococcal protein SdrE

Biochemical Journal ◽

10.1042/bcj20170132 ◽

2017 ◽

Vol 474 (11) ◽

pp. 1803-1806

Author(s):

Andrew B. Herr ◽

Alexander W. Thorman

Keyword(s):

Host Cell ◽

Immune Evasion ◽

Complement Activation ◽

Antimicrobial Agents ◽

Factor H ◽

Surface Proteins ◽

Host Cells ◽

Complement Factor ◽

Bacterial Killing ◽

Extracellular Matrix Components

The human immune system is responsible for identification and destruction of invader cells, such as the bacterial pathogen Staphylococcus aureus. In response, S. aureus brings to the fight a large number of virulence factors, including several that allow it to evade the host immune response. The staphylococcal surface protein SdrE was recently reported to bind to complement Factor H, an important regulator of complement activation. Factor H attaches to the surface of host cells to inhibit complement activation and amplification, preventing the destruction of the host cell. SdrE binding to Factor H allows S. aureus to mimic a host cell and reduces bacterial killing by granulocytes. In a new study published in Biochemical Journal, Zhang et al. describe crystal structures of SdrE and its complex with the C-terminal portion of Factor H. The structure of SdrE and its interaction with the Factor H peptide closely resemble a family of surface proteins that recognize extracellular matrix components such as fibrinogen. However, unbound SdrE forms a novel ‘Closed’ conformation with an occluded peptide-binding groove. These structures reveal a fascinating mechanism for immune evasion and provide a potential avenue for the development of novel antimicrobial agents to target SdrE.

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Identification of Staphylococcus aureus surface protein SdrE as a complement factor H-binding molecule

Molecular Immunology ◽

10.1016/j.molimm.2011.06.311 ◽

2011 ◽

Vol 48 (14) ◽

pp. 1695 ◽

Cited By ~ 1

Author(s):

K.M. Cunnion ◽

J.A. Sharp ◽

P.S. Hair ◽

C.G. Echague ◽

M.D. Ward ◽

...

Keyword(s):

Staphylococcus Aureus ◽

Surface Protein ◽

Factor H ◽

Complement Factor H ◽

Complement Factor

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Staphylococcus aureus Surface Protein SdrE Binds Complement Regulator Factor H as an Immune Evasion Tactic

PLoS ONE ◽

10.1371/journal.pone.0038407 ◽

2012 ◽

Vol 7 (5) ◽

pp. e38407 ◽

Cited By ~ 60

Author(s):

Julia A. Sharp ◽

Charlene G. Echague ◽

Pamela S. Hair ◽

Michael D. Ward ◽

Julius O. Nyalwidhe ◽

...

Keyword(s):

Staphylococcus Aureus ◽

Immune Evasion ◽

Surface Protein ◽

Factor H ◽

Complement Regulator

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Severe Congenital Thrombocytopenia Characterized by Decreased Platelet Sialylation and Moderate Complement Activation Caused by Novel Compound Heterozygous Variants in GNE

Frontiers in Immunology ◽

10.3389/fimmu.2021.777402 ◽

2021 ◽

Vol 12 ◽

Author(s):

Karolina I. Smolag ◽

Marcus Fager Ferrari ◽

Eva Zetterberg ◽

Eva Leinoe ◽

Torben Ek ◽

...

Keyword(s):

Sialic Acid ◽

Complement Activation ◽

Alternative Pathway ◽

Factor H ◽

Host Cells ◽

Lectin Pathway ◽

Compound Heterozygous ◽

Complement Factor ◽

Normal Platelet ◽

Compound Heterozygous Variants

BackgroundHereditary thrombocytopenias constitute a genetically heterogeneous cause of increased bleeding. We report a case of a 17-year-old boy suffering from severe macrothrombocytopenia throughout his life. Whole genome sequencing revealed the presence of two compound heterozygous variants in GNE encoding the enzyme UDP-N-acetyl-glucosamine-2-epimerase/N-acetylmannosamine kinase, crucial for sialic acid biosynthesis. Sialic acid is required for normal platelet life span, and biallelic variants in GNE have previously been associated with isolated macrothrombocytopenia. Furthermore, sialic acid constitutes a key ligand for complement factor H (FH), an important inhibitor of the complement system, protecting host cells from indiscriminate attack.MethodsSialic acid expression and FH binding to platelets and leukocytes was evaluated by flow cytometry. The binding of FH to erythrocytes was assessed indirectly by measuring the rate of complement mediated hemolysis. Complement activation was determined by measuring levels of C3bBbP (alternative pathway), C4d (classical/lectin pathway) and soluble terminal complement complex assays.ResultsThe proband exhibited markedly decreased expression of sialic acid on platelets and leukocytes. Consequently, the binding of FH was strongly reduced and moderate activation of the alternative and classical/lectin complement pathways was observed, together with an increased rate of erythrocyte lysis.ConclusionWe report two previously undescribed variants in GNE causing severe congenital macrothrombocytopenia in a compound heterozygous state, as a consequence of decreased platelet sialylation. The decreased sialylation of platelets, leukocytes and erythrocytes affects the binding of FH, leading to moderate complement activation and increased hemolysis.

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The N2N3 domains of ClfA, FnbpA, and FnbpB in Staphylococcus aureus bind to human complement factor H, and their antibodies enhance the bactericidal capability of human blood

The Journal of Biochemistry ◽

10.1093/jb/mvaa142 ◽

2020 ◽

Author(s):

Xinrui Mao ◽

Junghyun Kim ◽

QingFeng Zhang ◽

TingTing Jiang ◽

Dong Ho Ahn ◽

...

Keyword(s):

Staphylococcus Aureus ◽

Cell Wall ◽

Cell Surface ◽

Human Blood ◽

Bacterial Cell ◽

Regulatory Protein ◽

Factor H ◽

Complement Factor ◽

Bacterial Cell Surface ◽

Protein Factor

Abstract In the complement system, the opsonin C3b binds to the bacterial cell surface and mediates the opsonophagocytosis. However, the cell wall protein SdrE of Staphylococcus aureus inhibits the C3b activity by recruiting the complement regulatory protein factor H (fH). SdrE binds to fH via its N-terminal N2N3 domain, which are also found in six other staphylococcal cell wall proteins. In this study, we report that not only the N2N3 domain of SdrE but also those of ClfA, FnbpA, and FnbpB can bind to fH. When immobilized on a microplate, the N2N3 domains recruited fH and enhanced the factor I (fI)-mediated cleavage of C3b. When mixed with fH and S. aureus cells, the N2N3 domains inhibited the fH binding to S. aureus cells and reduced the fI-mediated C3b cleavage on the bacterial cell surface. The F(ab)′2 fragments of the rabbit N2N3 antibodies also inhibited the fH-binding to the S. aureus cell surface. When added to human blood, the N2N3 antibodies or the N2N3 domain proteins significantly increased the bactericidal activity. Based on these results, we conclude that, in S. aureus, not only SdrE but also ClfA, FnbpA, and FnbpB can contribute to the inhibition of C3b-mediated opsonophagocytosis.

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Complement factor H protects tumor cell-derived exosomes from complement-dependent lysis and phagocytosis

PLoS ONE ◽

10.1371/journal.pone.0252577 ◽

2021 ◽

Vol 16 (6) ◽

pp. e0252577

Author(s):

Ryan T. Bushey ◽

Elizabeth B. Gottlin ◽

Michael J. Campa ◽

Edward F. Patz

Keyword(s):

Tumor Cell ◽

Tumor Cells ◽

Cell Lines ◽

Alternative Pathway ◽

Metastatic Potential ◽

Factor H ◽

Host Cells ◽

Complement Factor H ◽

Complement Factor ◽

Classical Complement Pathway

Exosomes are a class of extracellular vesicles (EVs) that are mediators of normal intercellular communication, but exosomes are also used by tumor cells to promote oncogenesis and metastasis. Complement factor H (CFH) protects host cells from attack and destruction by the alternative pathway of complement-dependent cytotoxicity (CDC). Here we show that CFH can protect exosomes from complement-mediated lysis and phagocytosis. CFH was found to be associated with EVs from a variety of tumor cell lines as well as EVs isolated from the plasma of patients with metastatic non-small cell lung cancer. Higher levels of CFH-containing EVs correlated with higher metastatic potential of cell lines. GT103, a previously described antibody to CFH that preferentially causes CDC of tumor cells, was used to probe the susceptibility of tumor cell-derived exosomes to destruction. Exosomes were purified from EVs using CD63 beads. Incubation of GT103 with tumor cell-derived exosomes triggered exosome lysis primarily by the classical complement pathway as well as antibody-dependent exosome phagocytosis by macrophages. These results imply that GT103-mediated exosome destruction can be triggered by antibody Fc-C1q interaction (in the case of lysis), and antibody-Fc receptor interactions (in the case of phagocytosis). Thus, this work demonstrates CFH is expressed on tumor cell derived exosomes, can protect them from complement lysis and phagocytosis, and that an anti-CFH antibody can be used to target tumor-derived exosomes for exosome destruction via innate immune mechanisms. These findings suggest that a therapeutic CFH antibody has the potential to inhibit tumor progression and reduce metastasis promoted by exosomes.

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