Abstract
The sequence and blocking group of the amino-terminal 15 amino acids of rabbit trypsin-solubilized cytochrome b5 were determined by liquid secondary ion mass spectrometry (LSIMS) and tandem mass spectrometry (MS/MS). The molecular weights of peptides generated from aStaphylococcus aureus V8 protease digest of this protein were determined by LSIMS analysis and the two peptides containing the blocked amino-terminus were sequenced by tandem mass spectrometry to yield the sequence; N-acetyl-Ala-Ala-Glu-Ser-Asp-Lys-Asp-Val-Lys-Tyr-Tyr-Thr-Leu-Glu-Glu. Comparison of this sequence with a recently reported cDNA sequence (Dariushet al., 1988) indicates that Gln at position 3 is selectively deamidated, although no other discrepancies were found. Intact rabbit and bovine trypsin-solubilized cytochrome b5 were also analyzed by LSIMS on a high-field mass spectrometer equipped with a diode array detector. Mass measurement of the unresolved protonated molecular ion peak tops gave average molecular weights of 9462.2±2 and 9502.3±2 for bovine and rabbit trypsin-solubilized cytochrome b5, respectively. In both cases, these molecular weights correspond to a cytochrome b5 fragment consisting of amino acids Asp(7)-Arg(88). The average molecular weight for the rabbit amino-terminal-blocked form of trypsin-solubilized cytochrome b5 was found to be 10,144.5±2, which was consistent with the molecular weight predicted for the extended N-acetylated form (residues 1–88) of Mr 10,146.1.
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Gibson, B.W., Falick, A.M., Lipka, J.J. et al. Mass spectrometric analysis of rabbit and bovine trypsin-solubilized cytochrome b5. J Protein Chem 9, 695–703 (1990). https://doi.org/10.1007/BF01024764
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DOI: https://doi.org/10.1007/BF01024764