Establishment of a combined strategy of genetic and mass spectrometric analyses for characterizing hemoglobin mutations - An example of Hb Hoshida (beta 43Glu -> Gln)

Structural analysis of mutant hemoglobins has been efficiently accomplished by a consecutive mass spectrometric strategy: molecular mass measurement o f the protein to detect mutation and to determine the molecular mass change , and fragmentation analysis with collision-induced dissociation to determi ne the site and type of mutation. A flaw of this method is an inherent inab ility to detect a mutation associated with no or little change of the molec ular mass. In the present study, the strategy was improved by incorporating genetic analysis prior to mass spectrometry, which confirms the resulting amino acid change and searches for possible post-translational modification s. The method was applied to an unknown mutant, and elucidated a substituti on of glutamine for glutamic acid at position 43 of beta-globin subunit, th e mutation of Hb Hoshida. (C) 1999 Elsevier Science B.V. All rights reserve d.