Role of mass spectrometry in mapping strain variation and post-translational modifications of viral proteins.
Enzymatically derived fragments of the nucleocapsid protein from one strain (V4) of the paramyxovirus, New castle disease virus (NDV), have been aligned with the sequence deduced for a related strain (D26) by gene sequence analysis. This process involved extensive use of fast atom bombardment (FAB) mass spectrometry of unfractionated tryptic digests and fragments separated from tryptic or AspN protease digests by high-performance liquid chromatography (HPLC). Amino acid analysis and stepwise Edman degradation sequence analysis were used to complement FAB mass spectral data or as alternatives where no ions were produced by FAB. The nature of biosynthetic processing and blockage (acetylation) at the N-terminus of the protein were confirmed using collision-induced dissociation. Data obtained by direct analysis of the V4 nucleocapsid protein facilitated mapping of sequence variations within the nucleocapsid protein of the antigenically distinct WA2116 strain of NDV. Most of the WA2116 protein was mapped by FAB mass spectrometric analysis of HPLC fractions, thus amino acid analysis or stepwise sequence analysis were only required where FAB mass spectral data were inconclusive or indicated amino acid variations. This approach to comparison of NDV nucleocapsid proteins is proposed as a general strategy for mapping strain variation and post-translational modifications of viral proteins.
Gorman
J J
JJ
Commonwealth Scientific Industrial Research Organization, Australian Animal Health Laboratory, Victoria.
Corino
G L
GL
Shiell
B J
BJ
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D002213:Capsid / Q000032:analysis; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D002851:Chromatography, High Pressure Liquid; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D009522:Newcastle disease virus / Q000032:analysis / Q000378:metabolism; D010446:Peptide Fragments / Q000032:analysis; D011499:Protein Processing, Post-Translational; D013045:Species Specificity; D014357:Trypsin; D014764:Viral Proteins / Q000032:analysis* / Q000096:biosynthesis
D000596:Amino Acids; D010446:Peptide Fragments; D014764:Viral Proteins; D014357:Trypsin