scholarly journals Heavy chain-1 of inter-α-inhibitor has an integrin-like structure with immune regulatory activities

2019 ◽  
Author(s):  
David C. Briggs ◽  
Alexander W.W. Langford-Smith ◽  
Thomas A. Jowitt ◽  
Cay M. Kielty ◽  
Jan J. Enghild ◽  
...  
Keyword(s):  
Heavy Chain ◽  
Metal Ion ◽  
Chondroitin Sulphate ◽  
Binding Protein ◽  
Alternative Pathway ◽  
Complement C3 ◽  
Complement Factor ◽  
Heavy Chains ◽  
Covalent Complex ◽  
Adhesion Site

AbstractInter-α-inhibitor (IαI) is a proteoglycan essential for mammalian reproduction that also plays a less well-characterised role in inflammation. IαI is composed of 2 homologous ‘heavy chains’ (HC1 and HC2) covalently attached to chondroitin sulphate on the bikunin core protein. Prior to ovulation HCs are transferred onto the polysaccharide hyaluronan (HA), thereby stabilising a matrix that is required for fertilisation. Here we show that human HC1 has a structure similar to integrin β-chains and contains a functional MIDAS (metal ion-dependent adhesion site) motif that can mediate self-association of heavy chains, providing a mechanism for matrix crosslinking. Surprisingly, its interaction with RGD-containing integrin ligands, such as vitronectin and the latency-associated peptides of TGFβ, occurs in a MIDAS/cation-independent manner. However, HC1 utilises its MIDAS motif to bind to, and inhibit the cleavage of, complement C3, thus identifying it as a novel regulator of innate immunity through inhibition of the alternative pathway C3 convertase.AbbreviationsADPs, atomic displacement parameter; AUC, analytical ultracentrifugation; CMG2, capillary morphogenesis protein-2; COC, cumulus-oocyte complex; CS, chondroitin sulphate; FB, complement factor B; FnIII; fibronectin type III; HA, hyaluronan; HC, heavy chain; HC•HA, covalent complex of HC with HA; IαI, inter-α-inhibitor; ITGA, integrin α-chain; ITGB, integrin β-chain; LAP, latency associated peptide; LLC, large latent complex; LTBP, latent TGFβ binding protein; MIDAS, metal ion-dependent adhesion site; PαI, pre-α-inhibitor; PTX3, pentraxin-3; rHC1, recombinant HC1; SAXS, small-angle X-ray scattering; SHAP, serum-derived HA binding protein; SLC, small latent complex; TEM8, tumour endothelial marker-8; TGFβ, transforming factor β; TSG-6, tumour necrosis factor-stimulated gene-6; TSG-6•HC, covalent complex of TSG-6 and HC; vWFA domain, von Willebrand Factor A domain.

scholarly journals New functional and structural insights from updated mutational databases for complement factor H, Factor I, membrane cofactor protein and C3

2014 ◽  
Vol 34 (5) ◽  
Author(s):  
Elizabeth Rodriguez ◽  
Pavithra M. Rallapalli ◽  
Amy J. Osborne ◽  
Stephen J. Perkins
Keyword(s):  
Alternative Pathway ◽  
Factor H ◽  
Complement Factor H ◽  
Complement C3 ◽  
Complement Factor ◽  
Membrane Cofactor Protein ◽  
Complement Alternative Pathway ◽  
Factor I ◽  
Mutational Hotspots ◽  
Structural Insights

A new compilation of 324 mutations in four major proteins from the complement alternative pathway reveals mutational hotspots in factor H and complement C3, and less so in factor I and membrane cofactor protein. Their associations with function are discussed.

scholarly journals Inter-α-inhibitor heavy chain-1 has an integrin-like 3D structure mediating immune regulatory activities and matrix stabilization during ovulation

2020 ◽  
Vol 295 (16) ◽  
pp. 5278-5291 ◽  
Author(s):  
David C. Briggs ◽  
Alexander W. W. Langford-Smith ◽  
Holly L. Birchenough ◽  
Thomas A. Jowitt ◽  
Cay M. Kielty ◽  
...  
Keyword(s):  
Small Angle ◽  
Metal Ion ◽  
Alternative Pathway ◽  
3D Structure ◽  
Complement C3 ◽  
Dependent Manner ◽  
X Ray ◽  
X Ray Scattering ◽  
Matrix Stabilization ◽  
Ray Scattering

Inter-α-inhibitor is a proteoglycan essential for mammalian reproduction and also plays a less well-characterized role in inflammation. It comprises two homologous “heavy chains” (HC1 and HC2) covalently attached to chondroitin sulfate on the bikunin core protein. Before ovulation, HCs are transferred onto the polysaccharide hyaluronan (HA) to form covalent HC·HA complexes, thereby stabilizing an extracellular matrix around the oocyte required for fertilization. Additionally, such complexes form during inflammatory processes and mediate leukocyte adhesion in the synovial fluids of arthritis patients and protect against sepsis. Here using X-ray crystallography, we show that human HC1 has a structure similar to integrin β-chains, with a von Willebrand factor A domain containing a functional metal ion-dependent adhesion site (MIDAS) and an associated hybrid domain. A comparison of the WT protein and a variant with an impaired MIDAS (but otherwise structurally identical) by small-angle X-ray scattering and analytical ultracentrifugation revealed that HC1 self-associates in a cation-dependent manner, providing a mechanism for HC·HA cross-linking and matrix stabilization. Surprisingly, unlike integrins, HC1 interacted with RGD-containing ligands, such as fibronectin, vitronectin, and the latency-associated peptides of transforming growth factor β, in a MIDAS/cation-independent manner. However, HC1 utilizes its MIDAS motif to bind to and inhibit the cleavage of complement C3, and small-angle X-ray scattering–based modeling indicates that this occurs through the inhibition of the alternative pathway C3 convertase. These findings provide detailed structural and functional insights into HC1 as a regulator of innate immunity and further elucidate the role of HC·HA complexes in inflammation and ovulation.

Identity of the immunoglobulin heavy-chain-binding protein with the 78,000-dalton glucose-regulated protein and the role of posttranslational modifications in its binding function

1988 ◽  
Vol 8 (10) ◽  
pp. 4250-4256
Author(s):  
L M Hendershot ◽  
J Ting ◽  
A S Lee
Keyword(s):  
Heavy Chain ◽  
Posttranslational Modifications ◽  
Binding Protein ◽  
Fibroblast Cell ◽  
Immunoglobulin Heavy Chain ◽  
Lymphoid Cells ◽  
Temperature Sensitive ◽  
Heavy Chains ◽  
Binding Function ◽  
Glucose Regulated Protein

The 78,000-dalton glucose-regulated protein (GRP78) and the immunoglobulin heavy-chain-binding protein (BiP) were shown to be the same protein by NH2-terminal sequence comparison. Immunoprecipitation of GRP78-BiP induced by glucose starvation and a temperature-sensitive mutation in a hamster fibroblast cell line demonstrated the association of GRP78-BiP with other cellular proteins. In both fibroblasts and lymphoid cells, GRP78-BiP was found to label with 32Pi and [3H]adenosine. Phosphoamino acid analysis demonstrated that GRP78-BiP is phosphorylated on serine and threonine residues. Conditions which induce increased production of GRP78-BiP resulted in decreased incorporation of 32Pi and [3H]adenosine into GRP78-BiP. Furthermore, we report here that the phosphorylated form of BiP resides in the endoplasmic reticulum and that BiP which is associated with heavy chains is not phosphorylated or labeled with [3H]adenosine, whereas free BiP is. This suggests that posttranslational modifications may be important in regulating the synthesis and binding of BiP.

scholarly journals Post-translational processing of the inter-α-trypsin inhibitor in the human hepatoma HepG2 cell line

1994 ◽  
Vol 302 (2) ◽  
pp. 573-580 ◽  
Author(s):  
A Héron ◽  
J Bourguignon ◽  
A Callé ◽  
H Borghi ◽  
R Sesboüé ◽  
...  
Keyword(s):  
Heavy Chain ◽  
Trypsin Inhibitor ◽  
Hepg2 Cells ◽  
Chondroitin Sulphate ◽  
Brefeldin A ◽  
Human Hepatoma ◽  
Heavy Chains ◽  
Human Hepatoma Hepg2 Cells ◽  
Golgi Network ◽  
Hepatoma Hepg2

In human hepatoma HepG2 cells, the serum inter-alpha-trypsin inhibitor (ITI)-like protein is synthesized from two protein precursors, the heavy chain (H) H2 and the light chain (L). Both of them carry sulphate groups involved in the chondroitin sulphate glycosaminoglycan (GAG) linkage, as demonstrated by [35S]sulphate labelling, chondroitinase digestion and inhibition with beta-D-xyloside, an artificial GAG acceptor. While inhibition of N-glycosylation prevented neither the maturation nor the secretion of the ITI-related entities, brefeldin A induced the accumulation of H and L precursors in the cells, therefore blocking subsequent association and maturation of the precursors before their secretion. The enzyme system involved in the ester linkage between H and L chains is localized in the trans-Golgi network since no ITI-like protein could be obtained in the presence of monensin; instead free heavy-chain protein forms and bikunin were secreted in culture supernatants. The ITI-like protein synthesized by HepG2 cells is therefore composed of two heavy chains HC2 linked to two bikunin chains by chondroitin sulphate bridges, although the GAG linkage between HC2 chains is presumably different. Further, a different maturation route leading to restricted heavy-chain forms, Hm and Hd, could be shown.

scholarly journals Neisseria cinerea Expresses a Functional Factor H Binding Protein Which Is Recognized by Immune Responses Elicited by Meningococcal Vaccines

2017 ◽  
Vol 85 (10) ◽  
Author(s):  
Hayley Lavender ◽  
Katy Poncin ◽  
Christoph M. Tang
Keyword(s):  
Capsular Polysaccharide ◽  
Binding Protein ◽  
Alternative Pathway ◽  
Factor H ◽  
Recent Analysis ◽  
Complement Factor ◽  
Content Type ◽  
Commensal Flora ◽  
Commensal Species ◽  
Complement Regulator

ABSTRACT Neisseria meningitidis is a major cause of bacterial meningitis and sepsis worldwide. Capsular polysaccharide vaccines are available against meningococcal serogroups A, C, W, and Y. More recently two protein-based vaccines, Bexsero and Trumenba, against meningococcal serogroup B strains have been licensed; both vaccines contain meningococcal factor H binding protein (fHbp). fHbp is a surface-exposed lipoprotein that binds the negative complement regulator complement factor H (CFH), thereby inhibiting the alternative pathway of complement activation. Recent analysis of available genomes has indicated that some commensal Neisseria species also contain genes that potentially encode fHbp, although the functions of these genes and how immunization with fHbp-containing vaccines could affect the commensal flora have yet to be established. Here, we show that the commensal species Neisseria cinerea expresses functional fHbp on its surface and that it is responsible for recruitment of CFH by the bacterium. N. cinerea fHbp binds CFH with affinity similar to that of meningococcal fHbp and promotes survival of N. cinerea in human serum. We examined the potential impact of fHbp-containing vaccines on N. cinerea. We found that immunization with Bexsero elicits serum bactericidal activity against N. cinerea, which is primarily directed against fHbp. The shared function of fHbp in N. cinerea and N. meningitidis and cross-reactive responses elicited by Bexsero suggest that the introduction of fHbp-containing vaccines has the potential to affect carriage of N. cinerea and other commensal species.

scholarly journals COVID-19: a trigger for severe thrombotic microangiopathy in a patient with complement gene variant

2022 ◽  
Vol 0 (0) ◽  
Author(s):  
Larisa Pinte ◽  
Bogdan Marian Sorohan ◽  
Zoltán Prohászka ◽  
Mihaela Gherghiceanu ◽  
Cristian Băicuş
Keyword(s):  
Hemolytic Uremic Syndrome ◽  
Thrombotic Microangiopathy ◽  
Alternative Pathway ◽  
Atypical Hemolytic Uremic Syndrome ◽  
Factor H ◽  
Complement C3 ◽  
Uncontrolled Hypertension ◽  
Complement Factor ◽  
Uremic Syndrome ◽  
Complement Gene

Abstract The evidence regarding thrombotic microangiopathy (TMA) related to Coronavirus Infectious Disease 2019 (COVID-19) in patients with complement gene mutations as a cause of acute kidney injury (AKI) are limited. We presented a case of a 23-year-old male patient admitted with an asymptomatic form of COVID-19, but with uncontrolled hypertension and AKI. Kidney biopsy showed severe lesions of TMA. In evolution patient had persistent microangiopathic hemolytic anemia, decreased level of haptoglobin and increased LDH level. Decreased complement C3 level and the presence of schistocytes were found for the first time after biopsy. Kidney function progressively decreased and the patient remained hemodialysis dependent. Complement work-up showed a heterozygous variant with unknown significance in complement factor I (CFI) c.-13G>A, affecting the 5' UTR region of the gene. In addition, the patient was found to be heterozygous for the complement factor H (CFH) H3 haplotype (involving the rare alleles of c.-331C>T, Q672Q and E936D polymorphisms) reported as a risk factor of atypical hemolytic uremic syndrome. This case of AKI associated with severe TMA and secondary hemolytic uremic syndrome highlights the importance of genetic risk modifiers in the alternative pathway dysregulation of the complement in the setting of COVID-19, even in asymptomatic forms.

scholarly journals CFH and CFHR Copy Number Variations in C3 Glomerulopathy and Immune Complex-Mediated Membranoproliferative Glomerulonephritis

2021 ◽  
Vol 12 ◽  
Author(s):  
Rossella Piras ◽  
Matteo Breno ◽  
Elisabetta Valoti ◽  
Marta Alberti ◽  
Paraskevas Iatropoulos ◽  
...  
Keyword(s):  
Immune Complex ◽  
Single Molecule ◽  
Copy Number ◽  
Membranoproliferative Glomerulonephritis ◽  
Alternative Pathway ◽  
Copy Number Variations ◽  
Factor H ◽  
Complement C3 ◽  
Complement Factor ◽  
C3 Glomerulopathy

C3 Glomerulopathy (C3G) and Immune Complex-Mediated Membranoproliferative glomerulonephritis (IC-MPGN) are rare diseases characterized by glomerular deposition of C3 caused by dysregulation of the alternative pathway (AP) of complement. In approximately 20% of affected patients, dysregulation is driven by pathogenic variants in the two components of the AP C3 convertase, complement C3 (C3) and Factor B (CFB), or in complement Factor H (CFH) and Factor I (CFI), two genes that encode complement regulators. Copy number variations (CNVs) involving the CFH-related genes (CFHRs) that give rise to hybrid FHR proteins also have been described in a few C3G patients but not in IC-MPGN patients. In this study, we used multiplex ligation-dependent probe amplification (MLPA) to study the genomic architecture of the CFH-CFHR region and characterize CNVs in a large cohort of patients with C3G (n = 103) and IC-MPGN (n = 96) compared to healthy controls (n = 100). We identified new/rare CNVs resulting in structural variants (SVs) in 5 C3G and 2 IC-MPGN patients. Using long-read single molecule real-time sequencing (SMRT), we detected the breakpoints of three SVs. The identified SVs included: 1) a deletion of the entire CFH in one patient with IC-MPGN; 2) an increased number of CFHR4 copies in one IC-MPGN and three C3G patients; 3) a deletion from CFHR3-intron 3 to CFHR3-3′UTR (CFHR34–6Δ) that results in a FHR3-FHR1 hybrid protein in a C3G patient; and 4) a CFHR31–5-CFHR410 hybrid gene in a C3G patient. This work highlights the contribution of CFH-CFHR CNVs to the pathogenesis of both C3G and IC-MPGN.

scholarly journals Immunoglobulin heavy chain and binding protein complexes are dissociated in vivo by light chain addition.

1990 ◽  
Vol 111 (3) ◽  
pp. 829-837 ◽  
Author(s):  
L M Hendershot
Keyword(s):  
Heavy Chain ◽  
Light Chain ◽  
Secretory Pathway ◽  
Protein Transport ◽  
Protein Complexes ◽  
Binding Protein ◽  
Immunoglobulin Heavy Chain ◽  
Light Chains ◽  
Heavy Chains

Immunoglobulin heavy chain binding protein (BiP, GRP78) associates stably with the free, nonsecreted Ig heavy chains synthesized by Abelson virus transformed pre-B cell lines. In cells synthesizing both Ig heavy and light chains, the Ig subunits assemble rapidly and are secreted. Only incompletely assembled Ig molecules can be found bound to BiP in these cells. In addition to Ig heavy chains, a number of mutant and incompletely glycosylated transport-defective proteins are stably complexed with BiP. When normal proteins are examined for combination with BiP, only a small fraction of the intracellular pool of nascent, unfolded, or unassembled proteins can be found associated. It has been difficult to determine whether these BiP-associated molecules represent assembly intermediates which will be displaced from BiP and transported from the cell, or whether these are aberrant proteins that are ultimately degraded. In order for BiP to monitor and aid in normal protein transport, its association with these proteins must be reversible and the released proteins should be transport competent. In the studies described here, transient heterokaryons were formed between a myeloma line producing BiP-associated heavy chains and a myeloma line synthesizing the complementary light chain. Introduction of light chain synthesis resulted in assembly of prelabeled heavy chains with light chains, displacement of BiP from heavy chains, and secretion of Ig into the culture supernatant. These data demonstrate that BiP association can be reversible, with concordant release of transportable proteins. Thus, BiP can be considered a component of the exocytic secretory pathway, regulating the transport of both normal and abnormal proteins.

scholarly journals Access to the Complement Factor B Scissile Bond Is Facilitated by Association of Factor B with C3b Protein

2011 ◽  
Vol 286 (41) ◽  
pp. 35725-35732 ◽  
Author(s):  
Dennis E. Hourcade ◽  
Lynne M. Mitchell
Keyword(s):  
Conformational Changes ◽  
Metal Ion ◽  
Alternative Pathway ◽  
Salt Bridge ◽  
Single Amino Acid ◽  
Complement Factor ◽  
Salt Bridges ◽  
Complement Alternative Pathway ◽  
Factor B ◽  
Scissile Bond

Factor B is a zymogen that carries the catalytic site of the complement alternative pathway C3 convertase. During convertase assembly, factor B associates with C3b and Mg2+ forming a pro-convertase C3bB(Mg2+) that is cleaved at a single factor B site by factor D. In free factor B, a pair of salt bridges binds the Arg234 side chain to Glu446 and to Glu207, forming a double latch structure that sequesters the scissile bond (between Arg234 and Lys235) and minimizes its unproductive cleavage. It is unknown how the double latch is released in the pro-convertase. Here, we introduce single amino acid substitutions into factor B that preclude one or both of the Arg234 salt bridges, and we examine their impact on several different pro-convertase complexes. Our results indicate that loss of the Arg234-Glu446 salt bridge partially stabilizes C3bB(Mg2+). Loss of the Arg234-Glu207 salt bridge has lesser effects. We propose that when factor B first associates with C3b, it bears two intact Arg234 salt bridges. The complex rapidly dissociates unless the Arg234-Glu446 salt bridge is released whereupon conformational changes occur that activate the metal ion-dependent adhesion site and partially stabilize the complex. The remaining salt bridge is then released, exposing the scissile bond and permitting factor D cleavage.

scholarly journals Posttranslational association of immunoglobulin heavy chain binding protein with nascent heavy chains in nonsecreting and secreting hybridomas.

1986 ◽  
Vol 102 (5) ◽  
pp. 1558-1566 ◽  
Author(s):  
D G Bole ◽  
L M Hendershot ◽  
J F Kearney
Keyword(s):  
Monoclonal Antibody ◽  
Endoplasmic Reticulum ◽  
Cell Lines ◽  
Heavy Chain ◽  
Light Chain ◽  
B Lymphocyte ◽  
Binding Protein ◽  
Heavy Chains ◽  
Chain Synthesis ◽  
Ig Heavy Chain

A rat monoclonal antibody specific for immunoglobulin (Ig) heavy chain binding protein (BiP) has allowed the examination of the association of BiP with assembling Ig precursors in mouse B lymphocyte-derived cell lines. The anti-BiP monoclonal antibody immunoprecipitates BiP along with noncovalently associated Ig heavy chains. BiP is a component of the endoplasmic reticulum and binds free intracellular heavy chains in nonsecreting pre-B (mu+, L-) cell lines or incompletely assembled Ig precursors in (H+, L+) secreting hybridomas and myelomas. In the absence of light chain synthesis, heavy chains remain associated with BiP and are not secreted. The association of BiP with assembling Ig molecules in secreting hybridomas is transient and is restricted to the incompletely assembled molecules which are found in the endoplasmic reticulum. BiP loses affinity and disassociates with Ig molecules when polymerization with light chain is complete. We propose that the association of BiP with Ig heavy chain precursors is a novel posttranslational processing event occurring in the endoplasmic reticulum. The Ig heavy chains associated with BiP are not efficiently transported from the endoplasmic reticulum to the Golgi apparatus. Therefore, BiP may prevent the premature escape and eventual secretion of incompletely assembled Ig molecules.

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