Hot electron capture dissociation distinguishes leucine from isoleucine in a novel hemoglobin variant, Hb Askew, beta54(D5)Val-->Ile.

Population migration has led to the global dispersion of human hemoglobinopathies and has precipitated a need for their identification. An effective mass spectrometry-based procedure involves analysis of the intact alpha- and beta-globin chains to determine their mass, followed by location of the variant amino acid residue by direct analysis of the enzymatically digested chains and low-energy collision induced dissociation of the variant peptide. Using this procedure, a variant was identified as either beta54Val-->Leu or beta54Val-->Ile, since the amino acids leucine and isoleucine cannot be distinguished using low-energy collisions. Here, we describe how hot electron capture dissociation on a Fourier transform-ion cyclotron resonance mass spectrometer was used to distinguish isoleucine from leucine and identify the mutation as beta54(D5)Val-->Ile. This is a novel variant, and we have named it Hb Askew.