Hb Neuilly-sur-Marne, a new human hemoglobin variant with Ser-Asp-Leu inserted between

a86(F7) Leu and a87(F8) His: characterization by high-energy collision-induced dissociation liquid secondary ion mass spectrometry and low energy collision-induced dissociation tandem mass spectrometry in an ion trap fitted with a nanospray ionization source

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Eur. J. Mass Spectrom. 6, 205 - 211 (2000)

Hb Neuilly-sur-Marne, a new human hemoglobin variant with Ser-Asp-Leu inserted between a86(F7) Leu and a87(F8) His: characterization by high-energy collision-induced dissociation liquid secondary ion mass spectrometry and low energy collision-induced dissociation tandem mass spectrometry in an ion trap fitted with a nanospray ionization source

Danielle Promé^* and Jean-Claude Promé
Département de Chimie, 2R1, Université Paul Sabatier, 118 Route de Narbonne, 31062 Toulouse, France
Henri Wajcman, Jean Riou and Frédéric Galactéros
Inserm U468 et Département de Biochimie, Hôpital H. Mondor AP-HP, 94010 Crèteil, France
Nathalie Carte, Emanuelle Leize and Alain Vandorsselaer
Laboratoire de Spectrométrie de Masse, Université Louis Pasteur, 67084 Strasbourg, France

ABSTRACT:

Hemoglobin (Hb) Neuilly-sur-Marne is a new a-chain variant found during a systematic screening. Electrospray mass measurements showed the presence of an abnormal a-chain displaying a shift of u relative to the normal value. Tryptic cleavage of this chain and molecular weight determination of the peptides indicated that the 315 u shift was located into the aT-9 peptide, the molecular weight of which is higher than 3000 Da. High-energy collision spectra of MH⁺ ions generated by liquid secondary ion mass spectrometry from the normal and abnormal aT-9 afforded mainly amino-terminal containing ions. They indicated that these two peptides have an identical amino acid sequence from their 1st to 25th residues, the mass increase being thus located beyond this point. Too few ions were formed to establish reliably the sequence forward. It was hypothesized that this mass shift could result from a repeated sequence since the sum of the mass of the three residuesleucine, serine and aspartic acidpreceding position 25 is exactly 315 u. To get sequence information above position 25, decomposition of multicharged species was attempted. An ion trap fitted with a nanospray ionization source was used. It produced mainly triply- and quadruply-charged ions. Decomposition of the triply-charged ion afforded a series of singly-charged Y-ions in the expected region, giving a readily interpretable sequence. It confirmed the insertion of a Ser-Asp-Leu sequence above position 25. Surprisingly, decomposition of the quadruply-charged molecular ion gave too few ions to provide sequence information in the expected region. Spectra were dominated by some multicharged Y ions arising from cleavages close to the amino end. Tandem mass spectrometry experiments were performed on the abundant Y₃₀³⁺ ion and produced again a singly-charged Y ion series in the suitable domain which confirmed the above result. In Hb Neuilly-sur Marne this insertion of the Ser-Asp-Leu residues. between positions a-86 and a-87 is very likely due to a slipped strand mispairing mechanism.

Keywords: hemoglobin, insertion mechanism, LSIMS, high-energy collision, nanospray, ion trap, MSⁿ

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