Duffy blood group system - the frequency of Duffy antigen polymorphisms and novel mutations in the Polish population.

Duffy blood group genes are highly polymorphic with the distribution of alleles varying between different populations and ethnic groups. The aim of this study was to genotype Duffy blood group antigens and to establish FY alleles frequency in the Polish population and screen for novel FY gene mutations. Duffy phenotype and genotype frequencies analysis was based on studies of 596 persons.

[Duffy blood group antigens: structure, serological properties and function].

Duffy (Fy) blood group antigens are located on seven-transmembrane glycoprotein expressed on erythrocytes and endothelial cells, which acts as atypical chemokine receptor (ACKR1) and malarial receptor. The biological role of the Duffy glycoprotein has not been explained yet. It is suggested that Duffy protein modulate the intensity of the inflammatory response.

High-resolution melting analysis for genotyping Duffy blood group antigens.

Antigens of the Duffy (Fy) blood group are significant in medical transfusions since they may cause serious post-transfusion reactions and hemolytic disease of the fetus and newborn. Results of serotyping performed on donors with reduced or abolished erythrocyte Duffy expression may be misleading, since the Duffy antigen is also present on non-erythroid cells. In such cases only DNA-based genotyping may reveal the actual Duffy antigen status.

Studies of a murine monoclonal antibody directed against DARC: reappraisal of its specificity.

Duffy Antigen Receptor for Chemokines (DARC) plays multiple roles in human health as a blood group antigen, a receptor for chemokines and the only known receptor for Plasmodium vivax merozoites. It is the target of the murine anti-Fy6 monoclonal antibody 2C3 which binds to the first extracellular domain (ECD1), but exact nature of the recognized epitope was a subject of contradictory reports. Here, using a set of complex experiments which include expression of DARC with amino acid substitutions within the Fy6 epitope in E.

Molecular characterization of the Fy(a-b-) phenotype in a Polish family.

The Fy(a-b-) phenotype, very rare in Caucasians and defined by the homozygous FY(*)B-33 allele, is associated with the -33T>C mutation in the promoter region of the FY gene. The allele FY(*)X is correlated with weak expression of Fy(b) antigen due to 265C>T and 298G>A mutations in FY(*)B allele. The purpose of this study was molecular characterization of Fy blood group antigens in Fy(a-b-) members of a Polish family.

V(H)H (nanobody) directed against human glycophorin A: a tool for autologous red cell agglutination assays.

The preparation of a V(H)H (nanobody) named IH4 that recognizes human glycophorin A (GPA) is described. IH4 was isolated by screening a library prepared from the lymphocytes of a dromedary immunized by human blood transfusion. Phage display and panning against GPA as the immobilized antigen allowed isolating this V(H)H.

A single point mutation in the gene encoding Gb3/CD77 synthase causes a rare inherited polyagglutination syndrome.

Rare polyagglutinable NOR erythrocytes contain three unique globoside (Gb4Cer) derivatives, NOR1, NOR(int), and NOR2, in which Gal(α1-4), GalNAc(β1-3)Gal(α1-4), and Gal(α1-4)GalNAc(β1-3)Gal(α1-4), respectively, are linked to the terminal GalNAc residue of Gb4Cer. NOR1 and NOR2, which both terminate with a Gal(α1-4)GalNAc- sequence, react with anti-NOR antibodies commonly present in human sera. While searching for an enzyme responsible for the biosynthesis of Gal(α1-4)GalNAc, we identified a mutation in the A4GALT gene encoding Gb3/CD77 synthase (α1,4-galactosyltransferase).

One-step immunopurification and lectinochemical characterization of the Duffy atypical chemokine receptor from human erythrocytes.

Duffy antigen/receptor for chemokines (DARC) is a glycosylated seven-transmembrane protein acting as a blood group antigen, a chemokine binding protein and a receptor for Plasmodium vivax malaria parasite. It is present on erythrocytes and endothelial cells of postcapillary venules. The N-terminal extracellular domain of the Duffy glycoprotein carries Fy(a)/Fy(b) blood group antigens and Fy6 linear epitope recognized by monoclonal antibodies.

A recombinant dromedary antibody fragment (VHH or nanobody) directed against human Duffy antigen receptor for chemokines.

Fy blood group antigens are carried by the Duffy antigen receptor for chemokines (DARC), a red cells receptor for Plasmodium vivax broadly implicated in human health and diseases. Recombinant VHHs, or nanobodies, the smallest intact antigen binding fragment derivative from the heavy chain-only antibodies present in camelids, were prepared from a dromedary immunized against DARC N-terminal extracellular domain and selected for DARC binding. A described VHH, CA52, does recognize native DARC on cells.

Analysis of recombinant Duffy protein-linked N-glycans using lectins and glycosidases.

Duffy antigen is a glycosylated blood group protein acting as a malarial and chemokine receptor. Using glycosylation mutants we have previously demonstrated, that all three potential glycosylation sites of the Duffy antigen are occupied by N-linked oligosaccharide chains. In this study, wild-type Duffy glycoprotein and three mutants, each containing a single N-glycan, were used to characterize the oligosaccharide chains by lectin blotting and endoglycosidase digestion.

Construction of an agglutination tool: recombinant Fab fragments biotinylated in vitro.

The pComb3H vector system is used for constructing and panning recombinant antibody libraries. It allows for expression of monovalent Fab fragments, either on the surface of M13 phage, or in the form of soluble proteins secreted into the periplasmic space of bacteria. We constructed a modified pComb3H vector containing cDNA encoding for a 23-amino acid fragment of the Escherichia coli biotin carboxy carrier protein (BCCP), which is an acceptor sequence for biotinylation.

Murine monoclonal anti-s and other anti-glycophorin B antibodies resulting from immunizations with a GPB.s peptide.

Background: The blood group antigens S and s are defined by amino acids Met or Thr at position 29, respectively, on glycophorin B (GPB). Commercial anti-s reagents are expensive to produce because of the scarcity of human anti-s serum. Our aim was to develop hybridoma cell lines that secrete reagent-grade anti-s monoclonal antibodies (MoAbs) to supplement the supply of human anti-s reagents.

[Molecular background of the ABO blood group system].

The ABO human blood group system consists of A antigens, B antigens, and antibodies against these antigens. The antigenic determinants are synthesized in the Golgi apparatus by specific glycosyltransferases which transfer proper sugars to an oligosaccharide acceptor, called H antigen. N-acetylgalactosaminotransferase (transferase A) uses a UDP-GalNac donor to convert the H antigen to A antigen, whereas galactosyltransferase (transferase B) uses a UDP-galactose donor to convert the H antigen to B antigen.

[Interceptors:--"silent" chemokine receptors].

The physiological effect caused by chemokines is regulated by interactions with a group of rodopsin-like G protein-coupled receptors (GPCRs). These receptors share a number of common features: the polypeptide chain is a 7-transmembrane ?-helix (7 TMD motif) and the region involved in G-protein interaction (the DRYLAIV sequence) is located in the second transmembrane loop. So far, 19 chemokine receptors have been identified.

Mutational analysis of the N-glycosylation sites of Duffy antigen/receptor for chemokines.

The Duffy antigen/receptor for chemokines (DARC) is a seven-transmembrane glycoprotein carrying the Duffy (Fy) blood group antigen. The polypeptide chain of DARC contains two NSS motifs at positions 16 and 27 and one NDS motif at position 33 that represent canonical sequences for efficient N-glycosylation. To verify whether all of these three sites are occupied by a sugar chain, we generated mutants in which potential N-glycosylation sites (AsnXSer) were removed by replacement of serine by alanine.

[Chemokines--perspectives for using agents that block their action in therapy].

The chemokines are a large family of chemotactic cytokines, produced by tissue cells and leukocytes, which regulate leukocytes migration in inflammation and immunity. All the described human chemokines (approximately 50) transmit intracellular signals by binding and activating specific G protein-coupled receptors on the cell surfaces of their target cells. Chemokines appear to be involved in a variety of proinflammatory and autoimmune diseases, and this makes them and their receptors very attractive therapeutic targets.

The Fya, Fy6 and Fy3 epitopes of the Duffy blood group system recognized by new monoclonal antibodies: identification of a linear Fy3 epitope.

Four new anti-Duffy murine monoclonal antibodies (MAbs): two anti-Fy6 (MIMA-107 and MIMA-108), one anti-Fya (MIMA-19) and one anti-Fy3 (MIMA-29) were characterized. Identification of epitopes by means of synthetic peptides (Pepscan) showed that the anti-Fy6 reacted most strongly with peptides containing the sequence 19QLDFEDV25 of the Duffy glycoprotein, and less strongly with peptides containing LDFEDV (MIMA-107) or LDF only (MIMA-108). The anti-Fya recognized epitope 38DGDYGA43 containing the Gly42 residue, which defines the Fya blood group antigen.

Structure-function analysis of the extracellular domains of the Duffy antigen/receptor for chemokines: characterization of antibody and chemokine binding sites.

The Duffy antigen/receptor for chemokines (DARC), a seven-transmembrane glycoprotein carrying the Duffy (Fy) blood group, acts as a widely expressed promiscuous chemokine receptor. In a structure-function study, we analysed the binding of chemokines and anti-Fy monoclonal antibodies (mAbs) to K562 cells expressing 39 mutant forms of DARC with alanine substitutions spread out on the four extracellular domains (ECDs). Using synthetic peptides, we defined previously the Fy6 epitope (22-FEDVW-26), and we characterized the Fya epitope as the linear sequence 41-YGANLE-46.