Structural Characterization of Bovine Collectin‐43

Rothmann, Annette B.; Mortensen, Hanne D.; Holmskov, Uffe; Højrup, Peter

doi:10.1111/j.1432-1033.1997.t01-1-00630.x

Cited by 25 publications

(18 citation statements)

References 22 publications

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“…This model involves asymmetrical bonds, indicating a large flexibility of the polypeptide chains in the subunit. This pattern is identical to that determined for CL-43 (27). The N-terminal disulfide binding pattern has not previously been elucidated for any of the MBLs containing three cysteines, apart from the observation that the first of the three N-terminal cysteines seems to be responsible for the oligomerization, whereas the two other cysteines form intrasubunit bonds (25,26).…”

Section: Discussionsupporting

confidence: 62%

“…4). Again this is the result of the heterogeneity in the hydroxylation of Pro 21 and Pro 27 . As illustrated by Fig.…”

Section: Figmentioning

confidence: 99%

“…It is believed that the N-terminal cysteines and their bonding patterns are responsible for the variation in the oligomer structures of the collectins. Indeed, CL-43 and the nonoligomerizing form of rat MBL (MBL-C) show identical bonding patterns (22,27). The bonding patterns for the other collectins have not been solved.…”

Section: Fig 8 Western Blot Of the Cys To Ser Mutants Of Rhmblmentioning

confidence: 99%

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Characterization of the Oligomer Structure of Recombinant Human Mannan-binding Lectin

Jensen

Weilguny²,

Matthiesen³

et al. 2005

Journal of Biological Chemistry

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Mannan-binding lectin (MBL) belongs to a family of proteins called the collectins, which show large differences in their ultrastructures. These differences are believed to be determined by different N-terminal disulfide-bonding patterns. So far only the bonding pattern of two of the simple forms (recombinant rat MBL-C and bovine CL-43) have been determined. Recombinant MBL expressed in human cells was purified, and the structure of the N-terminal region was determined. Preliminary results on human plasma-derived MBL revealed high similarity to the recombinant protein. Here we report the structure of the N-terminal part of recombinant human MBL and present a model to explain the oligomerization pattern. Using a strategy of consecutive enzymatic digestions and matrix-assisted laser desorption ionization mass spectrometry, we succeeded in identifying a number of disulfide-linked peptides from the N-terminal cysteine-rich region. Based on these building blocks, we propose a model that can explain the various oligomeric forms found in purified MBL preparations. Furthermore, the model was challenged by the production of cysteine to serine mutants of the three N-terminally situated cysteines. The oligomerization patterns of these mutants support the proposed model. The model indicates that the polypeptide dimer is the basic unit in the oligomerization.Mannan-binding lectin (MBL) 1 is a serum protein that acts in innate immunity. It is a C-type lectin that recognizes and binds to specific sugars such as D-mannose and N-acetyl-Dglucosamine present on pathogen surfaces (1). The main functions in innate immunity are 1) the opsonin effect, where it binds to the surface of the pathogen and thereby enhances its clearance from the bloodstream (2) and 2) the ability to activate the complement cascade via the lectin pathway (3). Activation of the complement cascade requires binding of the MBL-associated serine proteases to the oligomeric forms of MBL (3-5) and leads to the formation of the membrane attack complex that perforates the cell membrane of the pathogen. The function of MBL is thus dependent on its oligomeric structure, since the small oligomer forms act as opsonins and the large oligomer forms activate complement (6 -8).The overall polypeptide structure of MBL is similar to that of the other collectins (surfactant protein A, surfactant protein D, conglutinin, CL-43, liver collectin 1, and CL-46). It includes a short, cysteine-rich N-terminal stretch (aa 1-21), a collagenlike region (aa 22-81) with one interruption (aa 43-44) that causes the collagen-like structure to bend, a neck region (aa 82-115), and a carbohydrate recognition domain (aa 116 -228) (9). This domain confers the carbohydrate specificity of MBL and is stabilized by two disulfide bonds (1). Due to the collagenlike domain, MBL forms homotrimers, designated the MBL subunit. The collagen-like structure is stabilized by the presence of hydroxyprolines and glycosylated hydroxylysines (10). The subunit structures assemble from the C to the N terminus. The neck...

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Section: Discussionsupporting

confidence: 62%

“…4). Again this is the result of the heterogeneity in the hydroxylation of Pro 21 and Pro 27 . As illustrated by Fig.…”

Section: Figmentioning

confidence: 99%

Section: Fig 8 Western Blot Of the Cys To Ser Mutants Of Rhmblmentioning

confidence: 99%

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Characterization of the Oligomer Structure of Recombinant Human Mannan-binding Lectin

Jensen

Weilguny²,

Matthiesen³

et al. 2005

Journal of Biological Chemistry

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“…These bovine serum collectins are structurally similar to SP-D; however, CL-43 is distinctive in that it only occurs as trimers in vivo and in vitro, whereas the others form dodecameric structures similar to SP-D (9,12,28). The bovine collectins also have distinctive monosaccharide binding preferences compared with SP-D.…”

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confidence: 89%

Viral aggregating and opsonizing activity in collectin trimers

Hartshorn

White

Tecle

et al. 2010

American Journal of Physiology-Lung Cellular and Molecular Phys

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Collectins are collagenous lectins present in blood, respiratory lining fluid, and other mucosal secretions that play important roles in innate defense against infection. The collectin, surfactant protein D (SP-D), limits infection by viruses and bacteria in the respiratory tract, eye, and female genital tract. Multimeric SP-D has strong antiviral activity and is a potent viral and bacterial agglutinin and opsonin; however, trimers composed of the neck and carbohydrate recognition domain (hSP-D-NCRD) of SP-D lack these activities. We now show that, in contrast, a trimeric neck and CRD construct of bovine serum collectin CL-46 induces aggregation of influenza A virus (IAV) and potently increases IAV uptake by neutrophils. CL-46-NCRD showed calcium-dependent and sugar-sensitive binding to both neutrophils and IAV. Replacement of specific residues of the CRD of human SP-D with those found in bovine serum collectins conferred opsonizing activity. The most effective substitution involved replacement of arginine 343 with valine (hSP-D-NCRD/R343V). hSP-D-NCRD/R343V greatly increased viral uptake by neutrophils and monocytes and also potentiated neutrophil respiratory burst responses. These effects were further increased by cross-linking of hSP-D-NCRD/R343V trimers with MAbs directed against areas of the hSP-D-NCRD not involved in viral binding. Unlike the wild-type human SP-D hSP-D-NCRD, hSP-D-NCRD/R343V also induced viral aggregation. These results indicate that collectins can act as opsonins for IAV even in the absence of the collagen domain or higher order multimerization. This may involve increased affinity of individual CRDs for glycoconjugates displayed on host cells or the viral envelope.

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“…Efforts are in progress to further characterize the abnormal cross-links; however, this will be a challenging task because both N-terminal cysteines normally participate in interchain bonds. It is likely that the native molecules contain both symmetrical (Cys 15 (56).…”

Section: Discussionmentioning

confidence: 99%

Myeloperoxidase-dependent Inactivation of Surfactant Protein D in Vitro and in Vivo

Crouch

Hirche

Shao

et al. 2010

Journal of Biological Chemistry

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Pulmonary surfactant protein D (SP-D)5 is a collagenous C-type lectin (collectin) that plays important, if not critical, roles in antimicrobial host defense, inflammatory regulation, and surfactant homeostasis (1-3). Although SP-D is primarily synthesized by respiratory epithelial cells and secreted in the alveolar spaces, it is also expressed at mucosal surfaces and other extrapulmonary sites, including the gastrointestinal and genitourinary tracts, where it could play similar roles in inflammatory and immune regulation (4).SP-D consists of one or more trimeric subunits, each with four structurally distinct domains (5, 6). The N-terminal domains mediate the association of subunits and contain conserved cysteine residues (Cys 15 and Cys 20 of the mature protein) that participate in intersubunit cross-links. By contrast, the C-terminal lectin domains mediate binding to microbial cell wall glycoconjugates, such as lipopolysaccharides (LPS), organic particulate antigens, nucleic acids, and specific cellular receptors (5,7,8). The intervening collagenous and neck domains maintain the trimeric structure of SP-D subunits and ensure an appropriate spatial distribution of the terminal lectin domains. Although the trimerization of CRDs is necessary for high affinity binding, higher order oligomerization of trimeric subunits is required for aggregation and bridging interactions of particulate ligands and effects on surfactant metabolism.Acute inflammation is characteristically accompanied by the recruitment and activation of neutrophils. SP-D can directly interact with neutrophils and modulate the antimicrobial functions of neutrophils in vitro (9, 10); it can also modestly enhance macrophage uptake of apoptotic neutrophils (11). SP-D-deficient mice show an exaggerated neutrophil response to viral and bacterial challenge (12,13). Recently, we demonstrated degradation of SP-D by neutrophil serine proteases (NSPs) in vitro and in vivo (14). Notably, all three granule-associated NSPs were able to cleave at specific sites within the functionally important lectin domain, abrogating carbohydrate recognition and bacterial aggregation. Together, these observations suggest a complex interplay between neutrophils and SP-D at sites of acute inflammation in vivo.In addition to proteases, neutrophil granules contain potent oxidant-generating enzymes, and recruited neutrophils are the major source of oxidants in the setting of acute inflammation

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Structural Characterization of Bovine Collectin‐43

Cited by 25 publications

References 22 publications

Characterization of the Oligomer Structure of Recombinant Human Mannan-binding Lectin

Characterization of the Oligomer Structure of Recombinant Human Mannan-binding Lectin

Viral aggregating and opsonizing activity in collectin trimers

Myeloperoxidase-dependent Inactivation of Surfactant Protein D in Vitro and in Vivo

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