scholarly journals cDNA cloning of a 30 kDa erythrocyte membrane protein associated with Rh (Rhesus)-blood-group-antigen expression

1990 ◽  
Vol 271 (3) ◽  
pp. 821-825 ◽  
Author(s):  
N D Avent ◽  
K Ridgwell ◽  
M J A Tanner ◽  
D J Anstee
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Membrane Protein ◽  
Blood Group ◽  
Erythrocyte Membrane ◽  
British Library ◽  
Cdna Clones ◽  
Blood Group Antigens ◽  
Hydrophobic Membrane ◽  
Pcr Products

The Rh-blood-group antigens (often described as Rhesus antigens) are associated with erythrocyte membrane proteins of approx. 30 kDa. We have determined the N-terminal 54 amino acid residues of the 30 kDa Rh D polypeptide (D30 polypeptide). We used primers based on these sequence data and the polymerase chain reaction (PCR) on human reticulocyte cDNA and genomic DNA to clone two types of PCR product of identical size. The two PCR products had related translated amino acid sequences between the 3′ ends of the primers, one of which was identical with that found for the D30 polypeptide. We designate the two related mRNA species which gave rise to the PCR products as Rh30A and Rh30B, the latter corresponding to the D30 polypeptide. We have isolated cDNA clones for the Rh30A protein which encode a hydrophobic membrane protein of 417 amino acids. The Rh30A protein has the same N-terminal 41 amino acids as the D30 polypeptide, but beyond this point the sequence differs, but is clearly related. The Rh30A protein probably corresponds to the R6A32 polypeptide, another member of the Rh 30 kDa family of proteins, which may carry the C/c and/or E/e antigens. Hydropathy analysis suggests that the Rh30A protein has up to 12 transmembrane domains. Three of these domains are bordered by a novel cysteine-containing motif, which might signal substitutions at these cysteine residues. Information which supplements this paper (amino-acid-sequence-analysis histograms) is reported in Supplementary Publication SUP 50160 (4 pages), which has been deposited at the British Library Document Supply Centre, Boston Spa, Wetherby, West Yorkshire LS23 7BQ, U.K., from whom copies can be obtained on the terms indicated in Biochem. J. (1990) 265, 5.

scholarly journals Isolation of cDNA clones for a 50 kDa glycoprotein of the human erythrocyte membrane associated with Rh (rhesus) blood-group antigen expression

1992 ◽  
Vol 287 (1) ◽  
pp. 223-228 ◽  
Author(s):  
K Ridgwell ◽  
N K Spurr ◽  
B Laguda ◽  
C MacGeoch ◽  
N D Avent ◽  
...  
Keyword(s):  
Amino Acid ◽  
Membrane Proteins ◽  
Amino Acid Sequence ◽  
Blood Group ◽  
Erythrocyte Membrane ◽  
Human Erythrocyte ◽  
Cdna Clones ◽  
Blood Group Antigens ◽  
Human Erythrocyte Membrane ◽  
Degenerate Primers

The Rh blood-group antigens are associated with human erythrocyte membrane proteins of approx. 30 kDa (the Rh30 polypeptides). Heterogeneously glycosylated membrane proteins of 50 and 45 kDa (the Rh50 glycoproteins) are coprecipitated with the Rh30 polypeptides on immunoprecipitation with anti-Rh-specific mono- and poly-clonal antibodies. We have isolated cDNA clones representing a member of the Rh50 glycoprotein family (the Rh50A glycoprotein). We used PCR with degenerate primers based on the N-terminal amino acid sequence of the Rh50 glycoproteins and human genomic DNA as a template and cloned and sequenced three types of PCR product of the expected size. Two of these products, Rh50A and Rh50B, gave the same translated amino acid sequence which corresponded to the expected Rh50 glycoprotein sequence but had only 75% DNA sequence similarity. The third product (Rh50C) contained a single base deletion, and the translated amino acid sequence contained an in-frame stop codon. We have isolated cDNA clones containing the full coding sequence of the Rh50A glycoprotein. This sequence predicts that it is a 409-amino acid N-glycosylated membrane protein with up to 12 transmembrane domains. The Rh50A glycoprotein shows clear similarity to the Rh30A protein in both amino acid sequence and predicted topology. Our results are consistent with the Rh30 and Rh50 groups of proteins being different subunits of an oligomeric complex which is likely to have a transport or channel function in the erythrocyte membrane. We mapped the Rh50A gene to human chromosome 6p21-qter, showing that genetic differences in the Rh30 rather than the Rh50 genes specify the major polymorphic forms of the Rh antigens.

scholarly journals Human erythrocyte acetylcholinesterase bears the Yta blood group antigen and is reduced or absent in the Yt(a-b-) phenotype

Blood ◽  
1993 ◽  
Vol 81 (3) ◽  
pp. 815-819 ◽  
Author(s):  
N Rao ◽  
CF Whitsett ◽  
SM Oxendine ◽  
MJ Telen
Keyword(s):  
Membrane Protein ◽  
Blood Group ◽  
Erythrocyte Membrane ◽  
Human Erythrocyte ◽  
Blood Cells ◽  
Phosphatidyl Inositol ◽  
Blood Group Antigen ◽  
Blood Group Antigens ◽  
Relationship Of ◽  
The Relationship

Abstract The Cartwright (Yt) blood group antigens have previously been shown likely to reside on a phosphatidylinositol-linked erythrocyte membrane protein. In this study, an unusual individual whose red blood cells (RBCs) were of the previously unreported Yt(a-b-) phenotype were used, along with normal Yt(a+) cells, to investigate serologically and biochemically the relationship of the Yta antigen to known phosphatidylinositol-linked erythrocyte proteins. Yt(a-b-) RBCs expressed normal amounts of various phosphatidyl-inositol-linked proteins except acetylcholinesterase. Further, human anti-Yta reacted with acetylcholinesterase in immunoprecipitation and immunoblotting studies. Thus, acetylcholinesterase is now identified as the protein bearing the Yt blood group antigens.

Human erythrocyte acetylcholinesterase bears the Yta blood group antigen and is reduced or absent in the Yt(a-b-) phenotype

Blood ◽  
1993 ◽  
Vol 81 (3) ◽  
pp. 815-819
Author(s):  
N Rao ◽  
CF Whitsett ◽  
SM Oxendine ◽  
MJ Telen
Keyword(s):  
Membrane Protein ◽  
Blood Group ◽  
Erythrocyte Membrane ◽  
Human Erythrocyte ◽  
Blood Cells ◽  
Phosphatidyl Inositol ◽  
Blood Group Antigen ◽  
Blood Group Antigens ◽  
Relationship Of ◽  
The Relationship

The Cartwright (Yt) blood group antigens have previously been shown likely to reside on a phosphatidylinositol-linked erythrocyte membrane protein. In this study, an unusual individual whose red blood cells (RBCs) were of the previously unreported Yt(a-b-) phenotype were used, along with normal Yt(a+) cells, to investigate serologically and biochemically the relationship of the Yta antigen to known phosphatidylinositol-linked erythrocyte proteins. Yt(a-b-) RBCs expressed normal amounts of various phosphatidyl-inositol-linked proteins except acetylcholinesterase. Further, human anti-Yta reacted with acetylcholinesterase in immunoprecipitation and immunoblotting studies. Thus, acetylcholinesterase is now identified as the protein bearing the Yt blood group antigens.

scholarly journals Protein-sequence studies on Rh-related polypeptides suggest the presence of at least two groups of proteins which associate in the human red-cell membrane

1988 ◽  
Vol 256 (3) ◽  
pp. 1043-1046 ◽  
Author(s):  
N D Avent ◽  
K Ridgwell ◽  
W J Mawby ◽  
M J Tanner ◽  
D J Anstee ◽  
...  
Keyword(s):  
Amino Acid ◽  
Blood Group ◽  
Protein Sequence ◽  
Amino Acid Sequences ◽  
Red Cells ◽  
British Library ◽  
Blood Group System ◽  
Terminal Amino Acid ◽  
Red Cell Membrane ◽  
Terminal Amino

The Rh D blood-group antigen forms part of a complex, involving several other polypeptides, that is deficient in the red cells of individuals who lack all the antigens of the Rh blood-group system (Rhnull red cells). These include components recognized by anti-(Rh D) antibodies and the murine monoclonal antibodies R6A and BRIC 125. We have carried out protein-sequence studies on the components immunoprecipitated by these antibodies. Anti-(Rh D) antibodies immunoprecipitate an Mr-30,000-32,000 polypeptide (the D30 polypeptide) and an Mr-45,000-100,000 glycoprotein (D50 polypeptide). Antibody R6A immunoprecipitates two glycoproteins of Mr 31,000-34,000 (R6A32 polypeptide) and Mr 35,000-52,000 (R6A45 polypeptide). The D30 and R6A32 polypeptides were found to have the same N-terminal amino acid sequences, showing that they are closely related proteins. The D50 polypeptide and the R6A45 polypeptide also had indistinguishable N-terminal amino acid sequences that differed from that of the D30 and R6A32 polypeptides. The putative N-terminal membrane-spanning segments of the two groups of proteins showed homology in their amino acid sequence, which may account for the association of each of the pairs of proteins during co-precipitation by the antibodies. Supplementary data related to the protein sequence have been deposited as Supplementary Publication SUP 50417 (6 pages) at the British Library Document Supply Centre, Boston Spa, Wetherby, West Yorkshire LS23 7BQ, U.K., from whom copies can be obtained on the terms indicated in Biochem. J. (1988) 249, 5.

Mouse protamine 2 is synthesized as a precursor whereas mouse protamine 1 is not

1987 ◽  
Vol 7 (6) ◽  
pp. 2173-2179
Author(s):  
P C Yelick ◽  
R Balhorn ◽  
P A Johnson ◽  
M Corzett ◽  
J A Mazrimas ◽  
...  
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Sequence Analysis ◽  
Nucleotide Sequence ◽  
Amino Acid Composition ◽  
Acid Composition ◽  
Nucleotide Sequence Analysis ◽  
Cdna Clones ◽  
Protamine 1 ◽  
Protamine 2

The nuclei of mouse spermatozoa contain two protamine variants, mouse protamine 1 (mP1) and mouse protamine 2 (mP2). The amino acid sequence predicted from mP1 cDNAs demonstrates that mP1 is a 50-amino-acid protein with strong homology to other mammalian P1 protamines. Nucleotide sequence analysis of independently isolated, overlapping cDNA clones indicated that mP2 is initially synthesized as a precursor protein which is subsequently processed into the spermatozoan form of mP2. The existence of the mP2 precursor was confirmed by amino acid composition and sequence analysis of the largest of a set of four basic proteins isolated from late-step spermatids whose synthesis is coincident with that of mP1. The sequence of the first 10 amino acids of this protein, mP2 precursor 1, exactly matches that predicted from the nucleotide sequence of cDNA and genomic mP2 clones. The amino acid composition of isolated mP2 precursor 1 very closely matches that predicted from the mP2 cDNA nucleotide sequence. Sequence analysis of the amino terminus of isolated mature mP2 identified the final processing point within the mP2 precursor. These studies demonstrated that mP2 is synthesized as a precursor containing 106 amino acids which is processed into the mature, 63-amino-acid form found in spermatozoa.

scholarly journals A novel O -linked glycan modulates Campylobacter jejuni major outer membrane protein-mediated adhesion to human histo-blood group antigens and chicken colonization

Open Biology ◽  
2014 ◽  
Vol 4 (1) ◽  
pp. 130202 ◽  
Author(s):  
Jafar Mahdavi ◽  
Necmettin Pirinccioglu ◽  
Neil J. Oldfield ◽  
Elisabet Carlsohn ◽  
Jeroen Stoof ◽  
...  
Keyword(s):  
Membrane Protein ◽  
Outer Membrane ◽  
Blood Group ◽  
Campylobacter Jejuni ◽  
Outer Membrane Protein ◽  
Ex Vivo ◽  
Binding Capacity ◽  
Major Outer Membrane Protein ◽  
Blood Group Antigens

Campylobacter jejuni is an important cause of human foodborne gastroenteritis; strategies to prevent infection are hampered by a poor understanding of the complex interactions between host and pathogen. Previous work showed that C. jejuni could bind human histo-blood group antigens (BgAgs) in vitro and that BgAgs could inhibit the binding of C. jejuni to human intestinal mucosa ex vivo. Here, the major flagella subunit protein (FlaA) and the major outer membrane protein (MOMP) were identified as BgAg-binding adhesins in C. jejuni NCTC11168 . Significantly, the MOMP was shown to be O- glycosylated at Thr 268 ; previously only flagellin proteins were known to be O- glycosylated in C. jejuni . Substitution of MOMP Thr 268 led to significantly reduced binding to BgAgs. The O- glycan moiety was characterized as Gal(β1–3)-GalNAc(β1–4)-GalNAc(β1–4)-GalNAcα1-Thr 268 ; modelling suggested that O- glycosylation has a notable effect on the conformation of MOMP and this modulates BgAg-binding capacity. Glycosylation of MOMP at Thr 268 promoted cell-to-cell binding, biofilm formation and adhesion to Caco-2 cells, and was required for the optimal colonization of chickens by C. jejuni , confirming the significance of this O- glycosylation in pathogenesis.

scholarly journals Human erythrocyte membrane sialoglycoprotein β. The cDNA sequence suggests the absence of a cleaved N-terminal signal sequence

1987 ◽  
Vol 243 (1) ◽  
pp. 277-280 ◽  
Author(s):  
S High ◽  
M J A Tanner
Keyword(s):  
Membrane Protein ◽  
Erythrocyte Membrane ◽  
Human Erythrocyte ◽  
Cdna Sequence ◽  
Signal Sequence ◽  
Human Kidney ◽  
Cdna Clones ◽  
Erythroid Cells ◽  
Coding Region ◽  
Human Erythrocyte Membrane

We have isolated cDNA clones corresponding to the human erythrocyte membrane sialoglycoprotein beta. The clones encompass the coding region for the protein, 120 residues of the 5′ non-coding region and the 3′ non-coding region. The cDNA sequence suggests that sialoglycoprotein beta is not translated with the cleaved N-terminal signal sequence usual in a membrane protein of this type. Sialoglycoprotein beta or a closely related homologue is present in human kidney as well as erythroid cells.

Predicting Structural and Serologic Modifications in Blood Group Antigens by Homology Modeling.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 451-451
Author(s):  
Connie M. Westhoff ◽  
Dwane E. Wylie
Keyword(s):  
Crystal Structure ◽  
Amino Acid ◽  
Homology Modeling ◽  
Blood Group ◽  
Amino Acid Change ◽  
Single Amino Acid ◽  
Side Chain ◽  
Blood Group Antigens ◽  
Aromatic Side Chain ◽  
Transport Channel

Abstract Homology modeling of blood group proteins offers the possibility of predicting the effect of amino acid changes on serologic phenotype and immunogenicity. The location of an amino acid change within known structural motifs, its phylogenetic conservation, and its proximity to known epitopes give insight into its potential effect on protein structure and, consequently, its clinical significance. We applied this approach to investigate the loss of membrane expression of the Dombrock blood group antigens in a patient with a single amino acid change and to investigate RhD alterations in weak D phenotypes. The Dombrock homology model was derived with rat ART2.2 crystal structure as template. For the RhD model, the crystal structure of the Rh-like-ammonia transporter from Nitrosomonas europaea was used. Protein alignment was derived with Clustal X, adjusted visually, and submitted to the Swiss Modeling server. Models were viewed with Deep View Swiss Pdb Viewer. The Dombrock null containes a Phe62Ser substitution. This Phe (F) residue is located in an FDDQY motif near the COOH terminus. This region of the protein also contains a HYYLT motif. These two motifs are highly conserved in the ART protein family and contribute several aromatic amino acids to this region of the molecule. Aromatic side chain interactions between these residues could contribute to the stability of the Do protein. In support, the distance in the ART2.2 crystal structure between Phe in FDDQY and His in HYYLT is 3.7 Å, which is the appropriate distance for aromatic side chain interactions. This is also the measured distance between these two residues in the Do model. Thus, protein modeling indicates that the Phe62Ser mutation disrupts important stacking interactions between Phe62 and His160. When amino acid changes causing weak D phenotypes were examined, some of those affecting expression of RhD were located near the vestigial transport channel. These include the Trp220Arg mutation (weak D Type 16). This Trp residue is part of the transport channel in Nitrosomonas and is conserved in Rh proteins of almost all species. Its role in maintaining Rh structure is indicated by the dramatic effect its modification has on protein and epitope expression. Additionally, Arg114Trp change (weak D Type 17), which is also near the channel, reduces D expression to only 66 antigen sites/cell. GlyXXXGly motifs stabilize interactions of adjacent alpha helices in membrane proteins. Evidence for a role in stabilization of RhD is revealed by the Gly282Asp mutation (weak D Type 15) which is part of such a motif. In addition, a D-epitope in loop 3 is near the 282Asp residue. Alteration of helical packing accompanied by epitope conformation could explain production of anti-D in patients with weak D Type 15. Homology modeling is an important tool for understanding the structure and serologic bases of blood group proteins and will continue to give important insight as more protein crystal structures become available.

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