Reference : HPA-1a phenotype-genotype discrepancy reveals a naturally occurring Arg93Gln substitu...

	Document type :	Scientific journals : Article
	Discipline(s) :	Life sciences : Biochemistry, biophysics & molecular biology
	To cite this reference:	http://hdl.handle.net/10993/8477

Title :	HPA-1a phenotype-genotype discrepancy reveals a naturally occurring Arg93Gln substitution in the platelet beta 3 integrin that disrupts the HPA-1a epitope.
Language :	English
Author, co-author :	Watkins, Nicholas A. [> >]
	Schaffner-Reckinger, Elisabeth [University of Luxembourg > Faculty of Science, Technology and Communication (FSTC) > Life Science Research Unit >]
	Allen, David L. [> >]
	Howkins, Graham J. [> >]
	Brons, Nicolaas H. C. [> >]
	Smith, Graham A. [> >]
	Metcalfe, Paul [> >]
	Murphy, Michael F. [> >]
	Kieffer, Nelly [> >]
	Ouwehand, Willem H. [> >]
Publication date :	2002
Journal title :	Blood
Volume :	99
Issue/season :	5
Pages :	1833-9
Peer reviewed :	Yes (verified by ORBilu)
Audience :	International
ISSN :	0006-4971
Country :	United States
Keywords :	[en] Amino Acid Substitution ; Animals ; Antibodies, Monoclonal/immunology ; Antigen-Antibody Reactions ; Antigens, CD/genetics ; Antigens, Human Platelet/genetics/immunology ; CHO Cells ; Cricetinae ; Epitopes, B-Lymphocyte/genetics/immunology ; Genotype ; Humans ; Integrin beta3 ; Mass Screening ; Phenotype ; Platelet Glycoprotein GPIIb-IIIa Complex/metabolism ; Platelet Membrane Glycoproteins/genetics ; Polymorphism, Single Nucleotide/genetics ; Sequence Analysis, DNA ; Transfection
Abstract :	[en] A single nucleotide polymorphism (SNP) at position 196 in the beta 3 integrin causes a Leu33Pro substitution in the mature protein. Alloimmunization against the beta 3Leu33 form (human platelet antigen [HPA]-1a, Pl(A1), Zw(a)) in patients who are beta 3Pro33 homozygous (HPA-1b1b, Pl(A2A2), Zw(bb)) causes neonatal alloimmune thrombocytopenia, posttransfusion purpura, or refractoriness to platelet transfusion. Studies with recombinant proteins have demonstrated that amino acids 1 to 66 and 288 to 490 of the beta 3 integrin contribute to HPA-1a epitope formation. In determining the HPA-1a status of more than 6000 donors, we identified a donor with an HPA-1a(weak) phenotype and an HPA-1a1b genotype. The platelets from this donor had normal levels of surface alpha IIb beta 3 but reacted only weakly with monoclonal and polyclonal anti-HPA-1a by whole blood enzyme-linked immunosorbent assay (ELISA), flow cytometry, and sandwich ELISA. We reasoned that an alteration in the primary nucleotide sequence of the beta 3Leu33 allele of this donor was disrupting the HPA-1a epitope. In agreement with this hypothesis, sequencing platelet RNA-derived alpha IIb and beta 3 cDNA identified a novel G/A SNP at position 376 of the beta 3 integrin that encodes for an Arg93Gln replacement in the beta 3Leu33 allele. Coexpression of the beta 3Leu33Gln93 encoding cDNA in Chinese hamster ovary cells with human alpha IIb cDNA showed that the surface-expressed alpha IIb beta 3 reacted normally with beta 3 integrin-specific monoclonal antibodies but only weakly with monoclonal anti-HPA-1a. Our results show that an Arg93Gln mutation in the beta 3Leu33 encoding allele disrupts the HPA-1a epitope, suggesting that Arg93 contributes to the formation of the HPA-1a B-cell epitope.
Permalink :	http://hdl.handle.net/10993/8477

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