HB Auckland [α87(F8)his→ASN]: A new Mutation of the Proximal Histidine Identified by Electrospray Mass Spectrometry

Abstract

Hb Auckland is a newly described unstable hemoglobin with a mutation of α87(F8)His→Asn. This substitution, involving the proximal histidine, does not lead to methemoglobinemia, but to instability and accelerated heme loss. The clinical picture is of a mild compensated hemolytic anemia. The presence of an abnormal hemoglobin was first demonstrated by the isopropanol stability test and confirmed by electrospray ionization mass spectrometry of total lysate. This showed that 14% of the α chains had a mass of 15,103.4 Da, i.e. 23 Da less than normal. Examination of tryptic digests showed an identical decrease in mass for peptide αT-9 (from 2,997.4 to 2,974.5 Da). Subdigestion with endoproteinase Asp-N located the 23 Da loss to residues α85-90, and further digestion with thermolysin identified the mutation as His→Asn at position 87 of the α chain. This was confirmed by sequence analysis of the peptide α85-90.