Structural Implications of Siglec-5-Mediated Sialoglycan Recognition

Zhuravleva, Marina A.; Trandem, Kathryn; Sun, Peter D.

doi:10.1016/j.jmb.2007.10.009

Cited by 66 publications

(79 citation statements)

References 43 publications

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“…We were interested in finding out which cysteine residue is involved in dimerization, and made a prediction based on three-dimensional structures using homology models and computational docking. Models based on the crystal structure of Siglec-5 (29) show that all six cysteines in V-set and C2set1 domains form disulfide bonds with each other either within a domain or between the domains (Fig. 5A).…”

Section: Resultsmentioning

confidence: 99%

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Studies on the Detection, Expression, Glycosylation, Dimerization, and Ligand Binding Properties of Mouse Siglec-E

Siddiqui

Schwarz

Springer

et al. 2017

Journal of Biological Chemistry

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Edited by Amanda F. Fosang CD33-related Siglecs are a family of proteins widely expressed on innate immune cells. Binding of sialylated glycans or other ligands triggers signals that inhibit or activate inflammation.Immunomodulation by Siglecs has been extensively studied, but relationships between structure and functions are poorly explored. Here we present new data relating to the structure and function of Siglec-E, the major CD33-related Siglec expressed on mouse neutrophils, monocytes, macrophages, and dendritic cells. We generated nine new rat monoclonal antibodies specific to mouse Siglec-E, with no cross-reactivity to Siglec-F. Although all antibodies detected Siglec-E on transfected human HEK-293T cells, only two reacted with mouse bone marrow neutrophils by flow cytometry and on spleen sections by immunohistochemistry. Moreover, whereas all antibodies recognized Siglec-E-Fc on immunoblots, binding was dependent on intact disulfide bonds and N-glycans, and only two antibodies recognized native Siglec-E within spleen lysates. Thus, we further investigated the impact of Siglec-E homodimerization. Homology-based structural modeling predicted a cysteine residue (Cys-298) in position to form a disulfide bridge between two Siglec-E polypeptides. Mutagenesis of Cys-298 confirmed its role in dimerization. In keeping with the high level of 9-O-acetylation found in mice, sialoglycan array studies indicate that this modification has complex effects on recognition by Siglec-E, in relationship to the underlying structures. However, we found no differences in phosphorylation or SHP-1 recruitment between dimeric and monomeric Siglec-E expressed on HEK293A cells. Phylogenomic analyses predicted that only some human and mouse Siglecs form disulfide-linked dimers. Notably, Siglec-9, the functionally equivalent human paralog of Siglec-E, occurs as a monomer. Two broad groups of Siglecs are identified based on their patterns of evolution. Sialoadhesin, CD22, myelin-associated glycoprotein (MAG) and Siglec-15 are highly conserved and show few changes among species (2). In contrast, CD33-related Siglecs evolve rapidly in mammals (2, 9). As there are not clear functional orthologs between primates and rodent Siglecs, these receptors are assigned an alphabetic letter in rodents and a number in primates. Siglec-E, the main CD33-related Siglec in mouse, was first discovered by a yeast two-hybrid screen using SHP-1 as a bait (10). Studies using a sheep polyclonal Siglec-E antibody specific for the ectodomain indicated that Siglec-E is expressed on neutrophils, macrophages, monocytes, dendritic cells, and a subset of Natural killer cells (11). The organ with the * This work was supported, in whole or in part, by National Institutes of Health Grants P01HL107150 and R01GM32373 (to A. V.). The authors declare that they have no conflicts of interest with the contents of this article. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health. □ S Th...

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Section: Resultsmentioning

confidence: 99%

“…Homology Modeling of Protein Structures-The three-dimensional structure of mouse Siglec-E was predicted based on an empirical structure of the V-set domain and first C2set domain of Siglec-5 (Protein Data Bank code 2ZG2) (29). Homology models of the V-set and C2set domains were generated in phyre2 (34), and visualized in PyMOL.…”

Section: Methodsmentioning

confidence: 99%

Studies on the Detection, Expression, Glycosylation, Dimerization, and Ligand Binding Properties of Mouse Siglec-E

Siddiqui

Schwarz

Springer

et al. 2017

Journal of Biological Chemistry

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“…5C) (29), further demonstrating that NKp30 uses a similar ligand binding site as that observed within the CD28 family of receptors. Interestingly, the F, G, and C strands also form the ligand binding sites in other I-type Ig-like receptors, such as ICAM and Siglec receptors (17,30,31). On the basis of these binding and mutational studies of NKp30 and B7-H6, their structural homology with PD-1 and CTLA-4 receptor ligand pairs, and the presence of a conserved structural recognition mechanism among members of the CD28 family receptors and their B7 ligands, we hypothesize that NKp30 may recognize other unidentified homologs of B7-H6 in a similar way to that seen in PD-1 and CTLA-4 ligand recognition (Fig.…”

Section: Resultsmentioning

confidence: 99%

Crystal structure of human natural cytotoxicity receptor NKp30 and identification of its ligand binding site

Joyce

Tran

Zhuravleva

et al. 2011

Proc. Natl. Acad. Sci. U.S.A.

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Natural killer (NK) cells are a group of innate immune cells that carry out continuous surveillance for the presence of virally infected or cancerous cells. The natural cytotoxicity receptor (NCR) NKp30 is critical for the elimination of a large group of tumor cell types. Although several ligands have been proposed for NKp30, the lack of a conserved structural feature among these ligands and their uncertain physiological relevance has contributed to confusion in the field and hampered a full understanding of the receptor. To gain insights into NKp30 ligand recognition, we have determined the crystal structure of the extracellular domain of human NKp30. The structure displays an I-type Ig-like fold structurally distinct from the other natural cytotoxicity receptors NKp44 and NKp46. Using cytolytic killing assays against a range of tumor cell lines and subsequent peptide epitope mapping of a NKp30 blocking antibody, we have identified a critical ligand binding region on NKp30 involving its F strand. Using different solution binding studies, we show that the N-terminal domain of B7-H6 is sufficient for NKp30 recognition. Mutations on NKp30 further confirm that residues in the vicinity of the F strand, including part of the C strand and the CD loop, affect binding to B7-H6. The structural comparison of NKp30 with CD28 family receptor and ligand complexes also supports the identified ligand binding site. This study provides insights into NKp30 ligand recognition and a framework for a potential family of unidentified ligands.

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“…To model the interaction between VAP-1 and Siglec-10, a structural model of the second C2-type domain of Siglec-10 was made based on the C2-set immunoglobulin domain of Siglec-5 (Protein Databank accession code 2zg2). 19 The reliability of the model is good because the immunoglobulin fold is in general known to be highly conserved and all the C2-type immunoglobulin domains found in CD33-related Siglecs share a high sequence similarity (supplemental Figure 1). Normally, the C2-type immunoglobulin fold is composed of 7 ␤-strands forming an anti-parallel ␤ barrel, but in Siglec-5 there are only 6 ␤-strands, ABE, and CFG.…”

Section: Siglec-10 Binds To the Enzymatic Groove Of Vap-1 And The Intmentioning

confidence: 98%

“…19 Ten models were generated with Modeller, 20 and the model with the lowest objective function was chosen. The sequence alignments were performed using MALIGN and MALFORM 21 within the Bodil visualization and modeling package.…”

Section: Modeling and Docking Simulationmentioning

confidence: 99%

Human Siglec-10 can bind to vascular adhesion protein-1 and serves as its substrate

Kivi

Elima

Aalto

et al. 2009

Blood

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Leukocytes migrate from the blood into areas of inflammation by interacting with various adhesion molecules on endothelial cells. Vascular adhesion protein-1 (VAP-1) is a glycoprotein expressed on inflamed endothelium where it plays a dual role: it is both an enzyme that oxidizes primary amines and an adhesin that is involved in leukocyte trafficking to sites of inflammation. Although VAP-1 was identified more than 15 years ago, the counterreceptor(s) for VAP-1 on leukocytes has remained unknown. Here we have identified Siglec-10 as a leukocyte ligand for VAP-1 using phage display screenings. The binding between Siglec-10 and VAP-1 was verified by different adhesion assays, and this interaction was also consistent with molecular modeling. Moreover, the interaction between Siglec-10 and VAP-1 led to increased hydrogen peroxide production, indicating that Siglec-10 serves as a substrate for VAP-1. Thus, the Siglec-10-VAP-1 interaction seems to mediate lymphocyte adhesion to endothelium and has the potential to modify the inflammatory microenvironment via the enzymatic end products. IntroductionMaintenance of an adequate immune defense is largely dependent on migration of circulating leukocytes from the vasculature into the surrounding tissue. Extravasation of leukocytes from the blood into the tissues is controlled by a multistep cascade, which involves numerous adhesion and signaling molecules. First, leukocytes tether and roll on the vascular endothelium, after which they adhere more strongly, arrest, and finally diapedese through the vessel wall. 1,2 One of the endothelial molecules involved in this cascade is a 180-kDa homodimeric glycoprotein, vascular adhesion protein-1 (VAP-1). 3 VAP-1 is mainly expressed on vascular endothelium, on smooth muscle cells, and on adipocytes. 4 It is rapidly translocated to the endothelial cell surface on inflammation, where it supports recruitment of leukocytes. 5,6 VAP-1 is involved in the rolling, firm adhesion, and transmigration phases of the extravasation cascade. Besides being an adhesin, VAP-1 is also an enzyme (semicarbazide sensitive amine oxidase, SSAO) and catalyzes oxidative deamination of a primary amine to an aldehyde with concomitant release of hydrogen peroxide and ammonium. 4 We and others have shown that VAP-1 supports leukocyte recruitment to sites of inflammation via both enzyme-activity-dependent and enzyme-activity-independent ways. Monoclonal anti-VAP-1 antibodies, which do not block the enzymatic activity of VAP-1, still block the binding of leukocytes to the endothelium in vitro and in vivo, and inhibitors of the enzymatic function of VAP-1 are able to effectively prevent the interaction between leukocytes and endothelium in vitro and in vivo. 7,8 Moreover, it has been recently shown that the end products generated by the catalytic activity of VAP-1, especially hydrogen peroxide, play a role in the regulation of other homing-associated molecules and thus enhance the inflammatory response. 9,10 Although VAP-1 has been known already for a long time and ...

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Structural Implications of Siglec-5-Mediated Sialoglycan Recognition

Cited by 66 publications

References 43 publications

Studies on the Detection, Expression, Glycosylation, Dimerization, and Ligand Binding Properties of Mouse Siglec-E

Studies on the Detection, Expression, Glycosylation, Dimerization, and Ligand Binding Properties of Mouse Siglec-E

Crystal structure of human natural cytotoxicity receptor NKp30 and identification of its ligand binding site

Human Siglec-10 can bind to vascular adhesion protein-1 and serves as its substrate

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