Molecular Configuration of Rh D Epitopes as Defined by Site-Directed Mutagenesis and Expression of Mutant Rh Constructs in K562 Erythroleukemia Cells
| Autor: | M. Scott, Neil D. Avent, Wendy Liu, D. Voak, Jeffrey W. Jones |
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| Rok vydání: | 1999 |
| Předmět: | |
| Zdroj: | Blood. 94:3986-3996 |
| ISSN: | 1528-0020 0006-4971 |
| DOI: | 10.1182/blood.v94.12.3986 |
| Popis: | The Rh D antigen is the most clinically important protein blood group antigen of the erythrocyte. It is expressed as a collection of at least 37 different epitopes. The external domains of the Rh D protein involved in epitope presentation have been predicted based on the analysis of variant Rh D protein structures inferred from their cDNA sequences and their D epitope expression. This analysis can never be absolute because (1) most partial D phenotypes involve multiple amino acid changes in the Rh D protein and (2) deficiency for 1 or more epitopes may be due to gross structural alteration in the variant Rh D protein structure. We report here the amino acid requirements for the majority of D epitopes. They have been defined by generating a series of novel Rh mutant constructs by mutagenesis using an Rh cE cDNA as template and mutagenic oligonucleotide primers. When transfected into K562 cells, the D epitope expression of the derived mutant clones was then assessed by flow cytometry. The introduction of 9 externally predicted Rh D-specific amino acids on the Rh cE protein was sufficient to express 80% of all tested D epitopes, whereas other clones expressed none. We concluded from our data that the D epitope expression is consistent with at least 6 different epitope clusters localized on external regions of the Rh D protein, most involving overlapping regions within external loops 3, 4, and 6. |
| Databáze: | OpenAIRE |
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