Characterization of hemoglobin bassett (α94Asp→Ala), a variant with very low oxygen affinity

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Abstract
Hemoglobin (Hb) Bassett, an abnormal Hb variant with a markedly reduced oxygen affinity, was discovered in a Caucasian (Anglo‐Saxon) male child who experienced episodes of cyanosis. Cation‐exchange and reversed‐phase (RP) high‐performance liquid chromatography (HPLC) showed that the patient has an abnormal Hb, with a mutation in the α‐globin. Tryptic peptide digest of the abnormal α‐globin with subsequent HPLC analysis revealed abnormal elution of the α‐T11 peptide. Further studies with Edman sequencing and electrospray mass spectrometry of tryptic peptide α‐T11, as well as structural analysis by X‐ray crystallography revealed an Asp→Ala substitution at the α94 (G1) position, a match for Hb Bassett. Detailed functional studies showed that this Hb variant had a markedly reduced oxygen affinity (P50 at pH 7.0 = 22 mmHg; Hb A P50 = 10.5 mmHg), reduced Bohr effect (−0.26 compared to − 0.54 in Hb A), and low subunit cooperativity (n = 1.4, compared to 2.6 in Hb A). X‐ray crystallography results explain the probable effects of the structural modification on the oxygen‐binding properties of this Hb variant. Am. J. Hematol. 77:268–276, 2004.