Rapid determination of sequence variations in actinidin isolated from Actinidia chinensis (var. Hayward) using fast atom bombardment mapping mass spectrometry and gas phase microsequencing.
A current limitation in the use of fast atom bombardment (FAB) mass spectrometric mapping of peptide mixtures, derived from enzymic digestion of proteins, is that most of the hydrophilic peptides are not observed. However, it has been demonstrated from previous work that esterification of the peptide mixture results in the detection of almost all peptides in FAB mass spectrometry. This strategy of FAB mapping was applied to the protein actinidin, isolated from an Italian variety of Actinidia chinensis. Two of the 12 tryptic peptides in FAB mass spectrometry did not exhibit molecular ions predicted from the known sequence of actinidin isolated from the New Zealand variety of A. chinensis. The two peptides were isolated by high-performance liquid chromatography, subjected to Staphylococcus aureus V8 protease digestion and sequenced by gas-phase microsequencing. Nine changes in amino acid composition were detected using the rapid and powerful combination of FAB mass spectrometric mapping and gas-phase microsequencing.
University Chemical Laboratory, University of Cambridge, U.K.
S
S
Naylor
S G
SG
Ang
D H
DH
Williams
C H
CH
Moore
K
K
Walsh
D000595:Amino Acid Sequence; D002849:Chromatography, Gas; D002851:Chromatography, High Pressure Liquid; D003546:Cysteine Endopeptidases / Q000032:analysis* / Q000302:isolation & purification; D005638:Fruit / Q000032:analysis*; D006868:Hydrolysis; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D010084:Oxidation-Reduction; D012697:Serine Endopeptidases; D013056:Spectrophotometry, Ultraviolet; D014357:Trypsin
D012697:Serine Endopeptidases; C028344:glutamyl endopeptidase; D014357:Trypsin; D003546:Cysteine Endopeptidases; C018745:actinidain