Electrospray mass spectrometric characterization of hemoglobin Q (Hb Q-India) and a double mutant hemoglobin S/D in clinical samples.

Abstract

Objectives: The clinical analysis of hemoglobin by ion exchange chromatography can result in ambiguities in identification of the nature of the globin chain present in patient samples. LC/ESI-MS provides rapid and precise determination of globin chain masses. Design and methods: Hemolysate of hemoglobin Q-India and hemoglobin S/D/F have been analyzed using ESI-MS. Tandem-MS has been used to establish mutation in α chain of hemoglobin Q. Results: The identification of hemoglobin Q-India is readily achieved by LC/ESI-MS, which establishes the presence of a mutant α chain differing in mass from normal α chain by 22 Da. The site of mutation has been identified by tandem-MS analysis of a tryptic fragment encompassing residues αV62-K90. LC/ESI-MS screening has also provide an example of simultaneous occurrence of mutant globin chains containing β6E → V (Hb S, sickle) and β121E→ Q (Hb D) variant. Expression of γ_G globin chain is also demonstrated in this sample. Conclusions: The site of mutation in hemoglobin Q-India is identified as α64D → H which differs from mutations α74D → H in Hb Q-Thailand and α75D → H in Hb Q-Iran. Mass spectrometric analysis of hemoglobins from a patient and her parents suggests inheritance of mutant β globin genes from both parents.