A Dutch Fay with Hb Atlanta [β75(E19)Leu → Pr0]

Abstract

We have investigated four members of a three-generation Dutch family for a suspected hemoglobinopathy. Chronic hemolysis and a moderate macrocytic normochromic anemia with slight morphological abnormalities of the red cells was observed in all four. Hemoglobin chain synthesis in vitro and separation of the globin chains by reversed phase high performance liquid chromatography revealed an abnormal p-globin species in addition to the normal α and β chains. The decreased amount of normal β-globin and the low amount of unidentified protein suggested an unstable β-globin variant. An abnormal band was detected by isoelectrofocusing. In one family member tested, the hemoglobin in an erythrocyte lysate had decreased heat stability. All carriers were positive in the isopropanol hemoglobin instability test. Treatment of erythrocytes with methylviolet gave rise tomicrogranular inclusions. Nucleotide sequencing of the polymerase chain reaction-amplified 0-globin gene revealed a heterozygous single base pair T → C mutation at codon 75, which changes the normal CTG codon for leucine to a CCG codon for proline. This variant has previously been identified as Hb Atlanta or β75(E19)Leu → Pro. The mutation creates a new Msp I restriction site, which was used to confirm the diagnosis in all four family members. A quantitative reverse transcriptase polymerase chain reaction procedure for determining the relative amounts of mRNA transcripts for the normal and abnormal globin chain showed a comparable stability for both transcripts.