Identification by Fast Atom Bombardment Mass Spectrometry of HB Indianapolis [β112(G14)CYS→ARG] in a Family from Naples, Italy

Abstract

We have characterized a P112 Arg hemoglobin in an individual from Naples, Italy, with minimal clinical problems.Blood tests revelead only slight reticulocytosis and hemoglobin instability. Furthermore, high value of alkali resistance tests for Hb F were observed. Isoelectricfocusing of globins showed the occurrence of a band migrating between the normal a and P globin chains. The fairly stable variant chain was purified by fast protein liquid chromatography. A mass map of the tryptic digest was obtained by fast atom bombardment mass spectrometry clearly showing that we were dealing with a p chain variant. However, the peptide 105-120 was missing and two new ones were present, i.e.: 105-112 and 113-120; we assumed these peptides to be generated because of the substitution of 112 Cys with an arginine residue. Further confirmation stemmed from the fast atom bombardment mass spectra of the tryptic digest submitted to a single Edman degradation step and to carboxypeptidase B further hydrolysis. The P-globin chain variant was thus mass mapped to an extent of about 98%. Such a variant, named

The Molecular Basis of -Thalassemia

“…This mutant was initially described as Hb Indianapolis (b112 Cys ! Arg) (Adams et al 1979), which was subsequently reported in two families (Spanish and Italian) (Baiget et al 1986;De Biasi et al 1988) in whom affected members had evidence of mild hemolytic anemia with 2% -4% reticulocytosis. In two patients heterozygous for Hb Terre Haute, globin-chain biosynthesis studies showed an a:non-a ratio of 1.0 in bone marrow erythroblasts compared with a ratio of 2.0 in peripheral blood reticulocytes.…”

A Rare Case of Variant Hemoglobin (Hb Yahata) Suspected Based on Inconsistent Plasma Glucose and HbA1c Levels

“…To date, three Hb variants with amino acid substitutions at the 112th position of the β chain have been reported: Hb Yahata, Hb Toranomon and Hb Indianapolis (11)(12)(13)(14)(15)(16)(17). In this case, Hb Yahata [β112 TGT (Cys) TAT (Tyr)] (heterozygote) was diagnosed.…”

“…Patients heterozygous for Hb Indianapolis [β 112 (G14) Cys Arg] reportedly have a very unstable variant with a phenotype similar to that of severe β-thalassemia (15). However, members of a Spanish family heterozygous for that variant were not anemic, despite exhibiting positive results on an in vitro test for unstable Hb (16). Hb Indianapolis was recently reported to cause moderate hemolytic anemia and renal damage in a Brazilian patient (17).…”

“…The amino acid at the 112th position of the β chain is an internal residue in the G helix that forms the central cavity and participates in the contact of α1β1 with Hb molecules. Therefore, the substitution of hydrophobic cysteine by another amino acid, such as arginine, tyrosine or tryptophan, likely results in inadequate adaptation and some degree of hemoglobin instability (10)(11)(12)(13)(14)(15)(16)(17).…”

IS IT DOMINANTLY INHERITED Β THALASSAEMIA OR JUST a Β‐CHAIN VARIANT THAT IS HIGHLY UNSTABLE?

“…It should be noted that the`real' Hb Indianapolis (b112 Cys ! Arg) has subsequently been reported in two families (Spanish and Italian) in whom affected members had evidence of mild haemolytic anaemia with 2±4% reticulocytosis (Baiget et al, 1986;De Biasi et al, 1988). Hb Indianapolis was mildly unstable and the b variant formed 41±43% of the total b-globin chains.…”