Analysis of Minor Hemoglobins by Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry

Post-translational Transformation of Methionine to Aspartate Is Catalyzed by Heme Iron and Driven by Peroxide

“…The mechanistic basis of Met 3 Asp conversion in HbF ␥V67M was examined using the following oxidative and nonoxidative conditions (experiments [1][2][3][4][5][6] O 2 (98% atom percent excess) as described above. 18 O labeling experiments were repeated with ferric HbF ␥V67M (condition 6) as described above.…”

“…The conversion to Met and Asp enhanced the spontaneous autoxidation of the mutants relative to wild-type HbA and human fetal Hb, and the levels of Asp were elevated with increasing levels of hydrogen peroxide (H 2 O 2 ). Using H 2 18 O 2 , we verified incorporation of 18 Human fetal hemoglobin (HbF) 3 and adult hemoglobin (HbA) are tetrameric heme proteins, which serve as the major transporters of oxygen (O 2 ) in fetal and adult life, respectively (1). These proteins consist of two globular ␣ subunits and two globular non-␣ subunits.…”

“…Post-translational modifications (PTMs) represent an important biological mechanism for modulating both protein function and increasing the overall functional diversity of the cellular proteome (1). These PTMs involve enzymatic and nonenzymatic modifications of amino acid side chains, including phosphorylation, methylation, acetylation, and glycosylation (2). Some of the commonly known post-translational changes in Hb, such as elevated levels of glycosylation in diabetes and carbamoylation due to elevated serum bicarbonate, are due to metabolic defects, and others are due to mutations that lead to covalent PTMs that can alter the stability of HbA or HbF in specific hemoglobinopathies (1).…”

Mass Spectrometry: A Tool for Enhanced Detection of Hemoglobin Variants

“…We performed cation-exchange HPLC (PolyCAT A column) according to (21 ) and RP HPLC as described in (22 ). Samples that were found with chromatographical methods to be abnormal were confirmed by IEF according to (23 ).…”

Detection of Compound Heterozygous of Hb Constant Spring and Hb Q-Thailand by Capillary Electrophoresis and High Performance Liquid Chromatography

“…However, it is difficult to diagnose the compound heterozygous of Hb CS and Hb Q-Thailand because Hb CS mRNA is unstable and it causes the rate of a-globin chain synthesis to decrease [8,9]. Moreover, some of the variants can co-elute and get masked on HPLC chromatogram and capillary electrophoresis (CE) electrophoregram [10,11]. A CE has proven to be superior to a HPLC in detecting Hb CS [12].…”